Reproductive styles of Osteoglossomorpha with

Reproductive styles of Osteoglossomorpha with emphasis on
Notopterus notopterus and Osteoglossum bicirrhosum
DISSERTATION
Zur Erlangung des akademischen Grades
Doctor rerum agriculturarum
(Dr. rer. agr)
Eingereicht an der
Landwirtschaftlich-Gärtnerischen Fakultät
der Humboldt-Universität zu Berlin
von
Honesty Yanwirsal
(M.Sc Fishery Science and Aquaculture)
Präsident der Humboldt-Universität zu Berlin
Prof. Dr. Jan-Hendrik Olbertz
Dekan der Landwirtschaflich-Gärtnerischen Fakultät
Prof. Dr. Dr. h. c. Frank Ellmer
Gutachter:
1. Prof. Dr. Frank Kirschbaum
2. Prof. Dr. Ralph Tiedemann
3. Dr. Peter Bartsch
Tag der mündlichen Prüfung: 16 Mai 2013
Dedicated to
Prof. Dr. Frank Kirschbaum
&
Dr. Peter Bartsch
Thousand words are never enough to express my gratitude.
It’s been a long journey, a precious and unforgettable experience in my life.
ii
LIST OF CONTENTS
LIST OF FIGURES ......................................................................................................... VI
LIST OF TABLES ........................................................................................................... IX
SUMMARY ..................................................................................................................... X
ZUSAMMENFASSUNG ............................................................................................... XII
1 INTRODUCTION ........................................................................................................1
1.1 General overview on Osteoglossomorpha ........................................................... 1
1.2 Reproduction in Osteoglossomorpha ..................................................................4
1.2.1 Influence of environmental factors on gonad development and
spawning season ...................................................................................... 4
1.2.1.1 Family Osteoglossidae .............................................................. 4
1.2.1.2 Family Notopteridae .................................................................4
1.2.1.3 Family Mormyridae ..................................................................5
1.2.1.4 Family Gymnarchidae ............................................................... 5
1.2.1.5 Family Pantodontidae ............................................................... 5
1.2.1.6 Family Hiodontidae ..................................................................5
1.2.2 Breeding behaviour .................................................................................6
1.2.2.1 Family Osteoglossidae .............................................................. 6
1.2.2.2 Family Notopteridae .................................................................7
1.2.2.3 Family Mormyridae ..................................................................7
1.2.2.4 Family Gymnarchidae ............................................................... 8
1.2.2.5 Family Pantodontidae ............................................................... 8
1.2.2.6 Family Hiodontidae ..................................................................8
1.2.3 Early ontogeny and objectives of the study ............................................8
2 MATERIALS AND METHODS ................................................................................10
2.1 Mormyridae .......................................................................................................10
2.1.1 Petrocephalus soudanensis ...................................................................10
2.1.2 Petrocephalus catostoma ......................................................................10
2.2 Pantodontidae ....................................................................................................11
2.2.1 Pantodon buchholzi...............................................................................11
2.3 Notopteridae ......................................................................................................13
2.3.1 Notopterus notopterus ...........................................................................13
2.3.1.1 Specimens and breeding tanks ................................................13
2.3.1.2 Rearing .................................................................................... 14
2.3.2 Xenomystus nigri ...................................................................................15
2.3.2.1 Specimens and breeding tanks ................................................15
2.4 Osteoglossidae ...................................................................................................16
2.4.1 Osteoglossum bicirrhosum ....................................................................16
2.4.1.1 Location and fishes .................................................................16
2.4.1.2 Collection of samples .............................................................. 16
iii
2.4.1.3 Rearing and documentation .................................................... 18
2.5 Variation and measurement of environmental parameters ................................ 19
2.6 Behaviours .........................................................................................................19
2.7 Maturity coefficient (MC) .................................................................................19
2.8 Hormonal treatment ........................................................................................... 20
2.9 General photography ......................................................................................... 20
2.10 Terminology ......................................................................................................20
3 RESULTS ...................................................................................................................21
3.1 Family Mormyridae ........................................................................................... 21
3.1.1 Petrocephalus soudanensis ...................................................................21
3.1.2 Petrocephalus catostoma ......................................................................23
3.2 Family Pantodontidae, Pantodon buchholzi...................................................... 26
3.3 Family Notopteridae .......................................................................................... 30
3.3.1 Xenomystus nigri ...................................................................................30
3.3.2 Notopterus notopterus ...........................................................................33
3.3.2.1 Reproduction ...........................................................................33
3.3.2.2 In situ condition of mature and immature (F1)
specimens ................................................................................39
3.3.2.3 External features of the egg .................................................... 40
3.3.2.4 Development ...........................................................................41
3.3.2.4.1 The embryonic period ....................................................... 41
3.3.2.4.1.1 The cleavage phase ....................................................... 41
3.3.2.4.1.2 The embryonic phase ...................................................43
3.3.2.4.1.3 The eleutheroembryonic phase .....................................49
3.3.2.4.2 The larval period ............................................................... 52
3.3.2.4.3 The juvenile period ........................................................... 54
3.3.2.4.4 The adult period ................................................................ 57
3.4 Osteoglossidae ...................................................................................................61
3.4.1 Osteoglossum bicirrhosum ....................................................................61
3.4.1.1 External morphology and in situ condition of
dissected gonads......................................................................61
3.4.1.2 Gonad of Silver Arowana and Blue Arowana ........................ 63
3.4.1.3 Egg of Silver Arowana............................................................ 64
3.4.1.4 Development ...........................................................................65
3.4.1.4.1 The embryonic period ....................................................... 65
3.4.1.4.1.1 The embryonic phase ...................................................65
3.4.1.4.1.2 The eleutheroembryonic phase .....................................71
3.4.1.4.2 The juvenile period ........................................................... 76
4 DISCUSSION .............................................................................................................81
4.1 Overview of environmental triggers of gonad development ............................. 81
4.1.1 Family Mormyridae ..............................................................................81
iv
4.2
4.3
4.4
4.1.2 Family Notopteridae .............................................................................81
4.1.3 Family Pantodontidae ...........................................................................82
4.1.4 Family Osteoglossidae ..........................................................................83
Modes of reproduction in Notopterus notopterus and
Osteoglossum bicirrhosum with comparison to other
osteoglossomorphs ............................................................................................ 83
4.2.1 Reproductive guilds ..............................................................................83
4.2.2 Spawning time....................................................................................... 84
4.2.3 Left gonad ............................................................................................. 84
4.2.4 Fractional spawner ................................................................................84
4.2.5 Parental care .......................................................................................... 85
4.2.6 Egg adhesiveness ..................................................................................85
4.2.7 Egg numbers per spawning ...................................................................86
4.2.8 Egg size .................................................................................................87
4.2.9 Hatching ................................................................................................ 87
4.2.10 Size of free embryos at hatching ........................................................... 88
4.2.11 Onset of exogenous feeding ..................................................................89
Characteristic of the egg envelope, eye pigmentation, melanophore
pattern and fins development in Notopterus notopterus and
Osteoglossum bicirrhosum with comparison to other
osteoglossomorphs ............................................................................................ 90
Comparison of periods and phases in development of
Notopterus notopterus and Osteoglossum bicirrhosum ....................................91
4.4.1 Periods ...................................................................................................92
4.4.2 Phases ....................................................................................................93
4.4.3 General comments.................................................................................93
5 CONCLUSION ...........................................................................................................95
6 REFERENCES ...........................................................................................................96
ACKNOWLEDGEMENTS .......................................................................................... 105
v
LIST OF FIGURES
Fig. 1. A male Petrocephalus soudanensis with a total length of 10.6 cm. .................... 10
Fig. 2. The experimental tank of Petrocephalus soudanensis .........................................11
Fig. 3. A male Petrocephalus catostoma with a total length of 10.9 cm.. ...................... 11
Fig. 4. A male Pantodon buchholzi with a total length of 9.6 cm ...................................12
Fig. 5. The experimental tank of the first group of Pantodon buchholzi. ....................... 12
Fig. 6. A female Notopterus notopterus with a total length of 23 cm.. ........................... 13
Fig. 7. Adult male and female Notopterus notopterus .................................................... 14
Fig. 8. Two experimental tanks of Notopterus notopterus.. ............................................14
Fig. 9. Adult female Xenomystus nigri with a total length of 10.7 cm............................ 15
Fig. 10. The genital papillae in Xenomystus nigri ........................................................... 15
Fig. 11. Collecting samples on the farm of Osteoglossum bicirrhosum. ........................ 17
Fig. 12. Separation of samples in Osteoglossum bicirrhosum. .......................................18
Fig. 13. Aquarium set up containing of Osteoglossum bicirrhosum. .............................. 18
Fig. 14. Course of environmental factors of Petrocephalus soudanensis ....................... 21
Fig. 15. Courtship behaviour in Petrocephalus soudanensis during the breeding
experiment ........................................................................................................22
Fig. 16. Illustration of the swimming movements of
Petrocephalus soudanensis ..............................................................................23
Fig. 17. Course of environmental factors in the breeding experiment group I of
Petrocephalus catostoma .................................................................................24
Fig. 18. Illustration of the position of the territories of
Petrocephalus catostoma during the day time .................................................24
Fig. 19. Course of environmental factors in the breeding experiment group II of
Petrocephalus catostoma .................................................................................25
Fig. 20. Course of environmental factors in the breeding experiment group I of
Pantodon buchholzi .......................................................................................... 27
Fig. 21. Course of environmental factors in the breeding experiment group II of
Pantodon buchholzi .......................................................................................... 27
Fig. 22. In situ condition of the gonads in Pantodon buchholzi. .....................................28
Fig. 23. Course of environmental factors in the breeding experiment group I of
Xenomystus nigri .............................................................................................. 30
Fig. 24. Course of environmental factors in the breeding experiment group II of
Xenomystus nigri .............................................................................................. 31
Fig. 25. In situ condition in Xenomystus nigri ................................................................ 32
Fig. 26. Course of environmental factors in breeding tank I of
Notopterus notopterus ...................................................................................... 34
vi
Fig. 27. Course of environmental factors in breeding tank II of
Notopterus notopterus. ..................................................................................... 35
Fig. 28. Two elements of courtship behaviour in Notopterus notopterus ....................... 36
Fig. 29. Intense removing gravels activity was found in breeding experiment II. ..........36
Fig. 30. Spawning sequence of Notopterus notopterus ...................................................38
Fig. 31. Preferred spawning sites of Notopterus notopterus ...........................................39
Fig. 32. Parental care of Notopterus notopterus performed by the male ........................ 39
Fig. 33. In situ condition in female Notopterus notopterus.............................................40
Fig. 34. Newly spawned eggs of Notopterus notopterus ................................................41
Fig. 35. Micropyle of Notopterus notopterus ..................................................................41
Fig. 36. Cleavage in Notopterus notopterus and blastulation .........................................44
Fig. 37. Continuation of epiboly and neurulation in Notopterus notopterus ..................45
Fig. 38. Embryonic development in Notopterus notopterus ...........................................46
Fig. 39. Hatching process and free embryonic in Notopterus notopterus ....................... 50
Fig. 40. Pectoral fin buds (pfb) in Notopterus notopterus ..............................................51
Fig. 41. Late embryonic and larval development in Notopterus notopterus ...................53
Fig. 42. Sequence of dorsal fin development in Notopterus notopterus ......................... 54
Fig. 43. Sequence of caudal fin development in Notopterus notopterus ........................ 55
Fig. 44. Two sequences of female’s genital papilla in Notopterus notopterus ...............55
Fig. 45. Complete development of female’s genital papilla in
Notopterus notopterus ...................................................................................... 56
Fig. 46. Juvenile transformation in Notopterus notopterus .............................................58
Fig. 47. Late juvenile to the maturation stage in Notopterus notopterus ........................ 59
Fig. 48. A female of Osteoglossum bicirrhosum............................................................. 61
Fig. 49. A female of Osteoglossum ferreirai (Blue Arowana) ........................................62
Fig. 50. Two ovaries of Osteoglossum bicirrhosum ....................................................... 63
Fig. 51. Freshly collected non-adhesive eggs of Osteoglossum bicirrhosum .................63
Fig. 52. First stages of the embryonic phase in Osteoglossum bicirrhosum ...................66
Fig. 53. Continuation of embryonic development in Osteoglossum bicirrhosum...........67
Fig. 54. Continuation of embryonic development in Osteoglossum bicirrhosum...........68
Fig. 55. Continuation of embryonic development in Osteoglossum bicirrhosum...........69
Fig. 56. Continuation of embryonic development in Osteoglossum bicirrhosum...........70
Fig. 57. Continuation of embryonic development of Osteoglossum bicirrhosum ..........71
Fig. 58. Pre-hatching and free embryonic development of
Osteoglossum bicirrhosum ...............................................................................72
Fig. 59. Free embryonic development of Osteoglossum bicirrhosum ............................ 73
vii
Fig. 60. Alteration of head structure of free embryo from stage 9 to stage 11 in
Osteoglossum bicirrhosum ...............................................................................74
Fig. 61. Starting the eleutheroembryonic phase of Osteoglossum bicirrhosum ..............75
Fig. 62. A mass of free embryos of Osteoglossum bicirrhosum. ....................................76
Fig. 63. Continuation of the eleutheroembryonic phase and starting point of the
juvenile stage in Osteoglossum bicirrhosum .................................................... 77
Fig. 64. Continuation of the juvenile sequences in Osteoglossum bicirrhosum .............79
Fig. 65. Comparison of time span of Notopterus notopterus and
Osteoglossum bicirrhosum ...............................................................................91
viii
LIST OF TABLES
Table 1. Measurement of total length and total weight of
Petrocephalus catostoma during the first breeding experiment ....................... 25
Table 2. Maturity coefficient (MC) of dissected Pantodon buchholzi at the end
of the experimental period, with total length (TL), total weight (TW),
and total gonad weight (TGW) data from all groups (I, II, CICombination I, CII-Combination II, CIII-Combination III, R-Rest) ...............29
Table 3. Maturity coefficient (MC) of dissected Xenomystus nigri at the end of
experimental period, with total length (TL), total weight (TW), and
total gonad weight (TGW) data from breeding group I and II ......................... 33
Table 4. Overview of spawning (Sp) events in breeding group I (2♀, 1♂) and
breeding group II (1♀, 1♂) with number of eggs, pH value and
temperature (T) per individual spawning ......................................................... 37
Table 5. Maturity coefficient (%) of dissected Notopterus notopterus, as
relating to total length (TL), total weight (TW), and total gonad
weight (TGW) data ........................................................................................... 40
Table 6. Overview of developmental stages of Notopterus notopterus (27 °C).
Determination of periods after Balon (1975) ...................................................60
Table 7. All visited farms of Osteoglossum bicirrhosum in Florencia and other
cities nearby together with the numbers of collected fresh eggs (♂1),
eggs containing embryo (♂2) and juveniles with yolk sac
(♂3 and ♂4) taken directly from male’s mouth ...............................................64
Table 8. Overview of developmental stages in Osteoglossum bicirrhosum
(28 °C), Determination after Balon (1975) ...................................................... 80
ix
SUMMARY
The Osteoglossomorpha represent a basal group of teleostean fish comprising taxa with
a mixture of both plesiomorphic and apomorphic characters of reproduction and
ontogenetic development. Concerning reproductive styles and ontogenetic development
of this group, there are still very limited data available so far. Some information on
different aspects of reproduction does exist. Most in depth studies are available for
mormyrids, but detailed descriptions and experimental data remain scarce in the other
groups as in Notopterus notopterus and Osteoglossum bicirrhosum.
This study will describe in detail for the first time the ontogenetic development of these
two species in laboratory-reared specimens. Breeding experiments aimed at potential
environmental triggers for gonad development or courtship behaviour of five species of
three families of the order Osteoglossiformes: Mormyridae (Petrocephalus soudanensis
and Petrocephalus catostoma), Pantodontidae (Pantodon buchholzi), and Notopteridae
(N. notopterus and Xenomystus nigri). For study of Osteoglossidae (O. bicirrhosum)
a-five-month field work took place in Colombia.
Only N. notopterus succeeded in the breeding experiment. Experimental data
demonstrated that the environmental factors decreasing conductivity, slight variation of
temperature, and water level have no influence on gonad development or courtship
behaviour in N. notopterus. Spawning occurs during day time at a temperature of
25-28 °C. Newly spawned 3.8–4 mm adhesive eggs are guarded by the male until
hatching. The egg envelope has external ridges, which are centred around the
micropyle. Hatching occurs within 168–204 hours. Exogenous feeding started on day 17
with a total length of 16.2 mm and yolk-sac remnants still present. The larval period
lasts until day 36. Dark brown stripes appear on the body as one of the characteristic
pigment patterns of juvenile N. notopterus at day 70 with a total length of 34 mm. The
genital papilla can macroscopically be recognized at day 80. Sexual maturity of
N. notopterus is first observed in 30-month old specimens with a total length of
275 mm.
For the first time, this study describes a method of successfully raising O. bicirrhosum
at 28 °C under laboratory conditions. The non-adhesive eggs measure 12 mm with a
transparent egg envelope. Hatching occurs around 162–166 hours and newly hatched
embryos measure 16 mm. Mixed feeding is observed at the age of 26 days, with the
juveniles reaching a total length of 38 mm. Around the age of 100 days and with a total
length of 125 mm, the juvenile is similar to an adult.
x
The embryonic period in O. bicirrhosum lasts longer than in N. notopterus. Actually
there is no larval period found in O. bicirrhosum. The embryonic period is directly
followed by the juvenile period and ontogeny can be characterized as direct
development. N. notopterus is classified as intermediate species in an interpretation at
reproductive strategies since they produce a higher number of medium-sized eggs and
show parental care.
xi
ZUSAMMENFASSUNG
Die Osteoglossomorpha stellen eine basale Gruppe der Teleostei dar mit einer
Mischung von plesiomorphen und apomorphen Merkmalen bezogen auf Reproduktion
und ontogenetische Entwicklung. Bezüglich reproduktiver Gilden und ontogenetischer
Entwicklung gibt es immer noch nur begrenzte Daten zu dieser Gruppe. Einige
Ergebnisse zu verschiedenen Aspekten der Reproduktion sind vorhanden. Der größte
Teil tiefer gehender Studien bezieht sich auf Mormyriden, detaillierte Beschreibungen
und experimentelle Daten sind kaum vorhanden bei den anderen Gruppen sowie bei
Notopterus notopterus und Osteoglossum bicirrhosum.
Im Rahmen dieser Untersuchung wird zum ersten Mal eine detaillierte Beschreibung
der ontogenetischen Entwicklung dieser beiden Arten auf der Basis von Laborzuchten
vorgestellt. Zuchtexperimente hatten zum Ziel, den möglichen Einfluss von
Umweltparametern auf Gonadenentwicklung und Balzverhalten von fünf Arten aus drei
Familien der Ordnung der Osteoglossiformes zu untersuchen: Mormyridae
(Petrocephalus soudanensis und P. catostoma), Pantodontidae (Pantodon buchholzi)
und Notopteridae (N. notopterus und Xenomystus nigri). Zum Studium der
Osteoglossidae (O. bicirrhosum) wurde eine fünfmonatige Felduntersuchung in
Kolumbien durchgeführt.
Nur bei N. notopterus gelangen Zuchtexperimente unter Laborbedingungen. Die
experimentellen Daten zeigten, dass die Umweltparameter abnehmende Leitfähigkeit,
Erhöhung des Wasserstandes und leichte Temperaturvariation keinen Einfluss auf
Gonadenentwicklung oder Balzverhalten bei N. notopterus hatten. Das Ablaichen
erfolgte zur Tageszeit bei 25-28 oC. Die frisch abgelegten, klebrigen Eier von 3.8-4 mm
Größe werden vom Männchen bis zum Schlupf bewacht. Die Eihülle besitzt äußere
Rillen, die ringförmig um die Mikropyle herum angeordnet sind. Das Schlüpfen erfolgt
im Alter von 168-204 Stunden. Exogene Nahrungsaufnahme begann am 17. Tag bei
einer Totallänge von 16.2 mm; zu diesem Zeitpunkt sind noch Reste des Dottersackes
vorhanden. Die Larval-Periode dauert bis zum 36. Tag. Dunkelbraune Querstreifen
erscheinen auf den Flanken als ein charakteristisches Juvenil-Farbmuster um Tag 70 bei
einer Gesamtlänge von 34 mm. Die Genitalpapille ist ab dem 80. Tag makroskopisch
sichtbar. Die Geschlechtsreife stellte sich bei 30 Monate alten Tieren ein bei einer
Totallänge von 275 mm.
In dieser Studie wird zum ersten Mal eine Methode zur erfolgreichen Aufzucht von
O. bicirrhosum unter Laborbedingungen bei 28 oC vorgestellt. Die nicht-klebrigen Eier
von 12 mm Größe besitzen eine transparente Eihülle. Das Schlüpfen erfolgt im Alter
xii
von 162-166 Stunden und die geschlüpften Embryonen haben eine Länge von 16 mm.
Exogene Nahrungsaufnahme beginnt bei gleichzeitigem Vorhandensein von einem
großen Dottersack bei einer Gesamtlänge der Juvenilen von 38 mm. Im Alter von etwa
100 Tagen und bei einer Gesamtlänge von ca. 125 mm besitzt der Juvenile schon die
Proportionen eines Adulten.
Die Embryonal-Periode bei O. bicirrhosum dauert länger als bei N. notopterus. Bei
O. bicirrhosum findet sich keine Larval-Periode; auf die Embryonal-Periode folgt sofort
die Juvenil-Periode: Die Ontogenese kann somit als direkte Entwicklung klassifiziert
werden. N. notopterus hingegen ist gekennzeichnet durch eine intermediäre
Entwicklung unter Bezug auf Reproduktionsstrategien da sie eine höhere Anzahl von
mittelgroßen
Eiern
produzieren
bei
gleichzeitiger
Brutpflege.
xiii
1 INTRODUCTION
1.1 General overview on Osteoglossomorpha
The Osteoglossomorpha or “bony tongues” comprise a group of morphologically and
biologically diverse primitive teleostean fish. The superorder was defined by
Greenwood et al. (1966). With a few exceptions, osteoglossomorphs have most of their
teeth located on the tongue (osteo, “bony”; glossid, “tongue”) and on the roof of the
mouth (or the parasphenoid) (Moyle, 2004). They also have a caudal fin with 16 or
fewer branched rays, no intermuscular bones in the back of the body (epipleurals),
cycloid scales with ornate microsculpturing, and an intestine that curls around to the left
side of the oesophagus rather than to the right as in most other bony fish (Nelson, 1972).
Although fossil evidence is sparse, osteoglossomorphs may have formed an important
element in the freshwater fauna of the world before the emergence of the
ostariophysans.
The Osteoglossomorpha contain two orders, the Osteoglossiformes and the
Hiodontiformes. Recent studies (see Li, 1994a; Li and Wilson, 1996a; Li et al., 1997)
support the concept of a sister-group relationship between the Hiodontiformes and the
Osteoglossiformes. The Osteoglossiformes consist of five families: the Osteoglossidae,
Notopteridae, Pantodontidae, Mormyridae, and Gymnarchidae. The Hiodontiformes on
the other hand are made up of only one single family, namely the Hiodontidae. The fish
of the family Osteoglossidae are found in South America, Africa, Asia, and Australia,
the Notopteridae in Africa and South-East Asia; the Mormyridae, Gymnarchidae and
Pantodontidae in Africa and the Hiodontidae solely in North America.
The relationship of the osteoglossomorphs to other fish groups is not fully defined,
presumably because they are such an ancient group. Greenwood (1973) suggested a
sister-group relationship between Osteoglossomorpha and Clupeomorpha, while
Patterson and Rosen (1977), Lauder and Liem (1983), and Nelson (1994) considered the
Osteoglossomorpha to be the most primitive living teleosts.
The seven species of the family Osteoglossidae are conspicuous members of their local
faunas, with heavy and elongate bodies. The dorsal and anal fins are long and placed on
the rear half of the body. Family characteristics also include: a scale-less head, a large
mouth usually turned upwards, pointed pectorals and small pelvic fins, a small or
reduced caudal and a mosaic-like pattern of large bony scales (Sterba, 1973;
Greenwood, 1975). Most species are predators and live in tropical rivers. All species of
the Osteoglossidae can breathe air through their lung-like swim bladders (Moyle, 2004).
1
The two species of Osteoglossum (O. bicirrhosum, O. ferreirai) exist in South America.
O. ferreirai (Black Arowana) can be exclusively found in the Negro River basin,
whereas O. bicirrhosum (Silver Arowana) inhabit the rest of the Amazon basin, the
Orinoco basin and various drainages of the Guyanas (Cala, 1973). The distribution of
these two species is based on different water types: O. ferreirai mainly occurs in acidic
black waters, while O. bicirrhosum is found in neutral or even slightly alkaline waters
(Saint-Paul et al., 2000). The Silver Arowana can reach up to 120 cm in length
(Lüling, 1976). O. bicirrhosum is known to be one of the most caught species in the
wild in South America, which has an impact on its population size. In Colombia,
O. bicirrhosum (Silver Arowana) is bred for the ornamental fish market. Australia is the
natural habitat of Scleropages leichhardtii, S. formosus and S. jardini. S. formosus can
be additionally found in South-East Asia and S. jardini in New Guinea. S. leichardtii
can reach up to 90 cm in length and 4 kg in weight, though it commonly measures
around 50 to 60 cm in length (Merrick et al., 1983). The Asian Arowana (S. formosus),
the most popular yet very expensive species on the aquarium trade market (Fernando et
al., 1997), is a large and attractive fish, reaching up to 7 kg in total weight and 100 cm
in total length (Alfred, 1964). The Saratoga, also called Spotted Barramundi, S. jardini,
was reported to attain a maximum length of more than 90 cm and maximum weight of
17.2 kg, though most specimens are 50 to 65 cm long (Anon, 1977). With a maximum
length of 300 cm, Arapaima gigas is one of the largest freshwater fish of South
America. The only member of the Osteoglossidae, that is not predatory but
planctivorous, is Heterotis niloticus from western Africa.
The family Notopteridae comprises four genera and 10 species with mostly long and
notably slender bodies. They are able to swim backwards equally well as forwards. The
dorsal fin is small and featherlike, so these fish are commonly referred to as “Feather
backs” or “Knife fish”. Knife fish have a long anal fin, which conjoins with the caudal
fin and closes behind the ventral fins. The mouth has many small teeth and the body is
covered with numerous tiny scales. The swim bladder is connected to the gut and is
used for breathing air. This swim bladder may have three functions: as a means for
aerial respiration, as an accessory auditory organ, and as an organ for sound production
(Greenwood, 1963). The knife fish live in stagnant backwaters and ponds.
Notopterus notopterus and the large growing species of the genus Chitala (C. chitala
and C. ornata) are fish from tropical South-East Asia (Moyle, 2004). Its natural habitats
are located in India, Pakistan, Burma, Thailand, the Malay Archipelago, the Philippines,
Cambodia, Vietnam, Laos, Bangladesh, and Myanmar (Day, 1889; Roberts, 1992).
N. notopterus
is
predominantly a
carnivorous
and
a
column-feeder
(Mustafa and Ahmed, 1979).
2
Xenomystus nigri occurs in Africa primarily in Nile, Chad, Niger, and Congo Basin
(Nelson, 2006). X. nigri can reach up to a maximum length of 22 cm and is adapted for
nocturnal life. The dorsal fin absents in this species. Some papers reported that X. nigri
possesses electroreceptors (Bullock and Northcutt, 1982; Braford, 1982). Other
notopterid species, Papyrocranus afer and P. congoensis also occur in Africa primarily
from Senegal to Nigeria and the Congo Basin (Nelson, 2006).
The family Mormyridae is a highly speciose group of African weakly electric fish.
According to some authors, it is considered as a separate teleostean order, the
Mormyriformes (Scott, 1973; Gosse, 1984; Nelson, 1994; Boden et al., 1997;
Alves-Gomes, 1999). They comprise around 200 species (Gosse, 1984) in about 15
genera of mormyrids. They are typically adapted for nocturnal life in large rivers and
lakes and are found in all tropical waters. There is a wide range of specialized
behaviours and a unique set of anatomical and physiological adaptations associated with
the electric sense (electroreception and electrogenesis). These features are believed to be
the success indicators of the radiation of this group. The electric system allows them to
move at night, to detect prey, and to communicate with each other. Weak electric
signals are produced by modified muscles in the caudal peduncle, and an electrical field
is set up around each fish (Moller, 1995). Larvae of mormyrids possess a distinct larval
electric organ (Kirschbaum, 1977). In addition to the significant electrical system in
mormyrids, these fish have a well-developed sense of hearing, which uses the swim
bladder to amplify sounds (Werns and Howland, 1976).
Gymnarchus niloticus of the monotypic family Gymnarchidae is closely related to the
mormyrids and also possesses an electric system for navigation and detecting prey
(Bass, 1982). G. niloticus has an elongate body, which it keeps rigid while swimming. It
lacks anal, caudal, and pelvic fins, but propels itself with its long dorsal fin, which
enables it to swim both forwards and backwards.
The family Pantodontidae is made up of only a single species, Pantodon buchholzi,
known as “Butterfly fish”. It is a small (10 cm), surface-feeding fish and an obligatory
air-breather (Schwartz, 1969). It is most remarkable for its specialized paired fins
(Moyle, 2004) and mostly distributed in the freshwaters of West Africa (Teugels, 1990)
and the Congo Basin. P. buchholzi has been considered as the sister group of the
Osteoglossidae (Greenwood et al., 1966; Nelson, 1969; Greenwood, 1973;
Taverne, 1979).
The family Hiodontidae has two extant species, the Mooneye (Hiodon tergisus) and the
Goldeye (H. alosoides). They are the most “normal”-looking fish in the entire
superorder, since they superficially resemble shad. Their most distinctive external
3
features are their large eyes with bright gold-silver irises in the Mooneye and gold irises
in the Goldeye. The Goldeye inhabits reservoirs, pools and strong currents in larger
rivers, and is tolerant of turbidity (Cross, 1967; Pflieger, 1997). This species is known
to perform a large-scale migration (Fernet and Smith, 1976). The Mooneye is generally
distributed throughout the Great Lakes, but is predominantly found in the waters of
Lake Erie in Ohio (Van Oosten, 1961). This species preferably inhabits clear waters and
mostly feeds in flowing waters, such as below dams (Trautman, 1957). Both species are
carnivorous, feeding on a wide variety of prey, but they are largely piscivorous as adults
(Scott and Crossman, 1973).
1.2 Reproduction in Osteoglossomorpha
1.2.1 Influence of environmental factors on gonad development and
spawning season
1.2.1.1 Family Osteoglossidae
In all six tropical species of osteoglossid the reproduction is indeed related to the high
and low water seasons. Argumedo (2005) reported about the breeding situation of
O. bicirrhosum in aquaculture, which occurs during rainy season starting in late
November and runs until the beginning of July. Between September and October
throughout rainy season S. leichardtii begins its courting behaviour (Merrick and
Green, 1982). Fish spawning in S. formosus occurs prior to the onset of the rainy season
(Scott and Fuller, 1976; Suleiman, 2003). In contrast, Rowley et al. (2000) stated that
the spawning season in S. formosus starts at the end of the dry season (March to April)
and lasts three months. As for Arapaima gigas (Lüling, 1964), reproduction mainly
appears to happen during the dry season (low water season).
1.2.1.2 Family Notopteridae
Notopterus notopterus in India mainly spawns during the rainy season with a maximum
Gonadosomato Index (GSI) of 6 occurring in June (Parameswaran and Sinha, 1966).
This statement was supported by Haniffa et al. (2004): a delay in spawning may occur
in India, when the monsoon arrives late. In Bangladesh, spawning in N. notopterus
commences with the beginning of the rainy season (Azadi et al, 1995). In Cambodia and
in the Ganges, N. notopterus spawns in the dry season (Mookerjee and Mazumdar,
1946; Chevey, 1941; Southwell and Prashad, 1919). Experimental data by Weitkamp
(2005) showed that gonad maturation in N. notopterus is independent of conductivity
and simulation of rain.
4
In other notopterids such as Papyrocronus afer spawning probably occurs in the rainy
season in the swamps of Gambia (Svensonn, 1933); and in the dry season in
Chitala chitala (Smith, 1933). Experimental data presented by Nyonje (2006) showed
that through the imitation of rain, increasing water level, and decrease of conductivity,
that gonad maturation in X. nigri is independent of three environmental factors
aforementioned.
1.2.1.3 Family Mormyridae
There are papers in mormyrids regarding the correlation of reproduction with the
alternation of dry and rainy seasons (Kirschbaum, 1975; 1987, 2006; Schugardt and
Kirschbaum, 2004, Kirschbaum and Schugardt, 2002; Kirschbaum et al., 2008;
Nguyen, 2011). This has been shown experimentally for Pollimyrus isidori
(Kirschbaum, 1975,
1987);
Mormyrus rume probocirostris
(Schugardt
and
Kirschbaum, 2004);
Hippopotamyrus pictus,
Campylomormyrus tamandua,
Campylomormyrus sp,
Pollimyrus adspersus,
Petrocephalus soudanensis
(Kirschbaum and Schugardt, 2001; Kirschbaum, 2006); and Paramormyrops of
the magnostipes-complex (Nguyen, 2011). The rainy season can be imitated (and the
gonad development induced) by solely decreasing conductivity.
1.2.1.4 Family Gymnarchidae
Spawning in Gymnarchus niloticus probably occurs during the rainy season
(Svensson, 1933) and towards the beginning of the flooding season in Senegal and in
the Gambia River (Hopkins, 1986).
1.2.1.5 Family Pantodontidae
So far there is only one reference available concerning the influence of environmental
factors on the reproduction of Pantodon buchholzi. Britz (2004) reported that egg
deposition in P. buchholzi occurs a few days after a drastic water change (50–80% of
tank volume), indicating therefore a correlation between reproduction and the rainy
season.
1.2.1.6 Family Hiodontidae
The two species of Hiodon: H. alosoides and H. tergisus spawn during summer
(dry season) and spring time (rainy season). According to Battles and Sprules (1960)
and Carlander (1969), peak spawning season in H. alosoides occurs at the beginning of
the dry season (late May to early July). In contrast, Cross and Collins (1995) mentioned
that spawning season in this species occurs during rainy season probably in March and
April.
5
In H. tergisus spawning occurs in early June and July during the dry season
(Fish, 1932). This stands in contrast to what Clark (1979) reported, namely that the peak
season of spawning in H. tergisus was during the rainy season around the middle of
March to April. Also Johnson (1951) stated that spawning in this species occurred in
May and April during rainy season. Since hiodontids inhabit temperate zone and are not
tropical fish, the influence of environmental factors on gonad maturation remains
unclear up to now.
1.2.2 Breeding behaviour
1.2.2.1 Family Osteoglossidae
The five Scleropages and Osteoglossum species are mouthbrooders and do not show
sexual dimorphism (Smith, 1931; Schaller and Dorn, 1971; Merick and Green, 1982;
Dawes et al., 1999; Scott and Fuller, 1976). Mouthbreeders in most of the mentioned
species in the Osteoglossidae are performed by the male (Argumedo, 2005; Merrick and
Green, 1982; Azuma, 1992; Brown, 1995; Schaller and Dorn, 1971; Takeshita, 1973;
Scott and Fuller, 1999; Dawes et al., 1999). Nevertheless, two papers also reported that
the female of Scleropages spp (Schaefer, 2010) and female of S. leichardtii (Merrick
and Schmida, 1984) are mouthbreeders.
In S. leichardtii spawning occurs at night in small ponds during spring time. The eggs of
S. leichardtii are around 10 mm in diameter (Lake and Midgley, 1970) and vary in
number from 30–130 per spawning (Allen et al., 2002). In S. formosus spawning occurs
in the afternoon (Azuma, 1992). Females of S. formosus produce large eggs around
30-100 (Scott and Fuller, 1976; Dawes et al., 1999) measuring 16-18 mm
(Azuma, 1992). During mouthbrooding some eggs maybe dead and embryos are
perhaps accidentally released from the parent’s mouth. This relates to the paper of
Rowley et al., (2000) who reported that a male of S. formosus in Cambodia may carry
up to 35 juveniles in his mouth.
In O. bicirrhosum, the onset of reproduction is indicated by the formation of a pair,
which gradually isolates from the group, establishes and defends a sector of the pond as
its territory (Argumedo, 2005). Wolfsheimer (1964) documented the large and yolky
eggs of O. bicirrhosum, which are 16 mm in size; eggs range from 40 (Ungar, 1993) up
to 150 (Maupin, 1967; Schaller and Dorn, 1971) per spawning. The eggs of
O. bicirrhosum after fertilization are reported to be 0.95 g in total weight and
approximate 13 mm in diameter (Aragao, 1981). Males of O. bicirrhosum and
O. ferreirai carry the offspring for three weeks and they do not feed at all during this
6
mouthbrooding phase (Schaller and Dorn, 1971). Sexual maturity starts for the first time
at an age of 30 to 36 months in O. ferreirai (Argumedo, 2009).
Heterotis niloticus (Budgett, 1901a; Svensson, 1933; Johnels, 1954; Daget, 1957) builds
nests and guards its eggs and young (Daget, 1957; Moreau, 1974). Arapaima gigas also
build nests and guard their eggs and young (Fontanele, 1948; Neves, 1998). In both
mentioned species, it is still unclear which sex guards the fry.
1.2.2.2 Family Notopteridae
Courtship and spawning activity in N. notopterus was observed during the day at a
temperature of 26-28 °C. This may last for about seven days (Friese, 1980). In contrast,
Pinxteren (1974) reported that spawning activity occurs during night and females lay
eggs on the ground and on stones in water temperature of 25 °C. The eggs are attached
to a substrate (Smith, 1933; Axelrod and Burgess, 1981; Friese, 1980). The 3–4 mm
eggs hatched within 11 days according to Axelrod and Burgess (1981), whereas Friese
(1980) mentioned that the eggs are 3.5 mm and hatched after six days at 28 °C.
Svensonn (1933) reported that newly spawned eggs of N. notopterus measured 4 mm.
Among Notopteridae, N. notopterus and the Chitala species perform parental care. The
male of N. notopterus and N. chitala guards the freshly spawned eggs (Pinxteren, 1974;
Smith, 1933; Axelrod and Burgess, 1981; Friese, 1980). X. nigri does not perform
parental care (Trittelvitz, 1986; Siraad, 1999).
1.2.2.3 Family Mormyridae
Sexual dimorphism becomes apparent at the anal fin in all investigated mormyrids, such
as Petrocephalus soudanensis (Kirschbaum, 2006); Pollimyrus isidori (Kirschbaum,
1987),
P. adspersus
(Kirschbaum
and
Schugardt,
2006),
and
Mormyrus rume probocirostris (Schugardt and Kirschbaum, 2004). Only Pollimyrus
(Kirschbaum, 1987; Diedhiou et al., 2007a) and Stomatorhinus (Heymer and Harder,
1975) show parental care in mormyrid species.
Spawning in mormyrids occurs mainly at night as seen in Campylomormyrus cassaicus,
Hippopotamyrus pictus, Mormyrus rume probocirostris, Pollimyrus isidori and
P. adspersus and Petrocephalus soudanensis (Kirschbaum, 1987, 2006; Kirschbaum
and Schugardt, 2002; Diedhiou et al., 2007a). Nguyen (2011) mentioned that spawning
after hormonal injection occurs at first during daylight in the fishes of
Paramormyrops magnostipes-complex. Spawning site preference exists in
Mormyrus rume probocirostris, Campylomormyrus cassaicus, Hippopotamyrus pictus
(Kirschbaum
and
Schugardt,
2004),
and
in
the
fishes
of
Paramormyrops magnostipes-complex (Nguyen, 2011; Arnegard et al., 2005).
7
1.2.2.4 Family Gymnarchidae
Gymnarchus niloticus builds large floating nests (Budgett, 1901a; Svensson, 1933);
containing around 1000 eggs with 10 mm in diameter (Budgett, 1901b). The embryos
hatched five days after spawning. Parental care is performed by the male.
1.2.2.5 Family Pantodontidae
The sex of Pantodon buchholzi can be determined by the anal fin and by the putative
copulatory organ (van Deurs and Lastein, 1973; van Deurs, 1973, 1975). This species
was assumed to possess an internal fertilization (Steche, 1915). Spawned eggs are
spherical, translucent and floating on the water surface due to large oil globules
(Britz, 2004). Eggs diameter measured 2.2–2.4 mm and free embryos hatched on the
third day at a temperature of 29 °C after egg deposition (Britz, 2004). This single
species exhibits no parental care (Mohn, 1976a; Britz, 2004).
1.2.2.6 Family Hiodontidae
Males of Hiodon aloisoides and H. tergisus have an anal fin with a convex anterior of
anal fin, while in females the anal fin is slightly concave (Scott and Crossman, 1973;
Hilton, 2002). Both species Hiodon alosoides (Battle and Sprules, 1960) and H. tergisus
(Synder and Douglas, 1978) do not exhibit parental care.
Sexual maturity in H. alosoides may occur as early as age 1 year (Synder and Douglas,
1978). Spawning is assumed to take place at night (Scott and Crossman, 1973). Eggs
deposited on rocks (Scott and Crossman, 1973) and gravel (Balon, 1975a). Eggs are
spherical, buoyant, non-adhesive with a single large oil globule, measuring 4 mm in
diameter (Fish, 1932). Hatching occurs in about 14 weeks (Scott and Crossman, 1973).
Sexual maturity in H. tergisus usually reached in third or fourth year at 228-280 mm
(Van Oosten, 1961). Eggs deposited over rocks and gravel (Balon, 1975b). Females
produce around 3000-7.700 eggs in Tennessee River (Wallus and Buchanan, 1989).
Fertilized eggs are spherical, non-adhesive and buoyant (Balon, 1975b).
1.2.3 Early ontogeny and objectives of the study
There are a number of papers concerning the early development in Osteoglossomorpha.
Argumedo (2005) has written a handbook on commercial breeding in aquaculture of
Osteoglossum bicirrhosum including a staging of development. Wolfsheimer (1964)
and Ungar (1993) reported briefly on the development of O. bicirrhosum. In general,
information on the early ontogeny of O. bicirrhosum is still very scarce.
8
Regarding notopterids, a recent study by Srivastava et al. (2012) described the
embryonic and larval development in fixed specimens of Notopterus notopterus.
However a huge gap remains concerning the detailed description of the egg and the egg
envelope, early cleavage, larval and juvenile development.
Regarding mormyrid species, the ontogeny of several species has been described, e.g. in
Hyperopisus bebe (Johnels, 1954), Pollimyrus adspersus (Kirschbaum, 1987),
Mormyrus rume proboscirostris (Kirschbaum and Schugardt, 1995; Schugardt and
Kirschbaum, 1996, 2004, 2006), Campylomormyrus tamandua (Schugardt and
Kirschbaum, 1998), Hippopotamyrus pictus (Kirschbaum and Schugardt, 2002b), and
Petrocephalus soudanensis (Kirschbaum, 2006). Yet only Diedhiou et al., (2007b)
documented for the first time in detail the ontogenetic development in the mormyrid
Pollimyrus isidori and proposed a staging system.
The early ontogenetic development in Gymnarchus niloticus has been described by
Budgett (1901), Assheton (1907) and Svensson (1933). However detailed information
on the early ontogeny of G. niloticus remains limited.
In Pantodon buchholzi, Britz (2004) described the newly spawned egg, its structure, the
newly hatched embryo and the larval development. He also summarized characteristics
of reproductive styles in 16 additional species of osteoglossomorphs.
In hiodontid species, Battle and Sprules (1960) published a description of the free
embryo stage and the larval stage in Hiodon alosoides. Synder and Douglas (1978)
presented a very short description of the free embryonic stage in H. tergisus. A
summary of morphometric data for early life phases of H. tergisus is described by
Wallus and Buchanan (1989).
This study will fill some gaps on early ontogenetic development and will present for the
first time a detailed ontogeny of Notopterus notopterus (Notopteridae) and
Osteoglossum bicirrhosum (Osteoglossidae) together with experimental data of two
mormyrid
species
Petrocephalus soudanensis
and
P. catostoma,
and
Pantodon buchholzi (Pantodontidae) and Xenomystus nigri (Notopteridae). Therefore,
the objectives of the present study are: 1) to present a detailed description of ontogenetic
development in N. notopterus and O. bicirrhosum, and 2) to describe and to evaluate the
characteristic differences between these two species: the substrate spawner
N. notopterus and the mouth breeder O. bicirrhosum. The study aims at a more detailed
comparison with other osteoglossomorphs in view of elucidating a bit more the
heterogeneous systematic distribution of different reproductive styles among
Osteoglossomorpha.
9
2 MATERIALS AND METHODS
2.1 Mormyridae
2.1.1 Petrocephalus soudanensis
The four specimens (3♂, 1♀) of Petrocephalus soudanensis (Fig. 1), which were used
for the first breeding experiment, were already present at the Humboldt University
before the start of this project. They were kept in a 155 x 62 x 50 cm tank (500–600 l)
equipped with a biofilter and PVC tubes as hiding places (Fig. 2). Conductivity was
decreased by continuously adding deionised water of a conductivity of 25 µS/cm.
Temperature and conductivity were measured daily by using a conductivity meter from
WTW Tetracon 325. The fish were fed with Chaoborus sp. twice a day.
Fig. 1. A male Petrocephalus soudanensis with a total length of 10.6 cm [a]. [b] The base line of the anal
fin (arrowhead) of the male is concave and has longer fin rays than [c] the female’s, whose base line of
the anal fin is straight. Scale bar = 3 cm.
2.1.2 Petrocephalus catostoma
Seven specimens (6♂, 1♀) of Petrocephalus catostoma (Fig. 3) were donated by
Prof. Dr. B. Kramer. The fish originated from the Namibia-East Caprivi upper Zambezi
River at Katima Mulilo, West Africa. The fish were kept in tanks similar to those of
10
P. soudanensis and the breeding experiment was performed in a similar way. During the
experiment, all tubes in the breeding tank were completely infested with planarians. The
tubes were used as hiding places. Therefore it was necessary to wash out the tank with
hot water twice a week.
Fig. 2. The experimental tank of Petrocephalus soudanensis equipped with plastic tubes (x) covered by a
plastic sheet (arrow).
Fig. 3. A male Petrocephalus catostoma with a total length of 10.9 cm. Scale bar = 3 cm.
2.2 Pantodontidae
2.2.1 Pantodon buchholzi
All specimens of the Butterfly fish Pantodon buchholzi (Fig. 4), originating from the
Niger, West Africa, were purchased from a whole sale dealer (Aquarium Glaser in
Rodgau, Frankfurt am Main). The first group of Pantodon (n=16) arrived on 5 February
2009, consisting of 11 males and 5 females. Total length ranged from 75-107 mm and
total weight from 3.74–9.93 g. The second group arrived on 6 May 2009, comprising 10
fish (9♀, 1♂), their total length ranging from 77–105 mm and their total weight from
11
2.45–6.94 g. The third group (n=100) was purchased on 10 December 2010. Due to
illness, 85 specimens died in the tank a week upon arrival. Only 15 specimens (5♀,
10♂) survived.
The sex of a butterfly fish can be clearly determined by the shape of its anal fin. The
female’s fin has a straight line at the rear edge. The male anal fin on the other hand is
notched or indented and almost looks like two fins (Fig. 4a, b), with a short and straight
posterior and long and filamentous anterior part (Vriends, 1978). They were fed on
crickets (Acheta domesticus) twice a day, ad libidum.
Fig. 4. A male Pantodon buchholzi with a total length of 9.6 cm [a]. The anal fin of a male [b] is notched
and a female’s is straight [c]. Scale bar = 2 cm.
Fig. 5. The experimental tank of the first group of Pantodon buchholzi equipped with black polythene
shreds (arrow) as plant imitation. The three small plastic tubes (arrowhead) were placed on the surface of
the water to prevent the crickets, which were fed to the fish, from leaving the tank.
12
The three groups of fish were put into three separate tanks: group one in a
200 x 60 x 50 cm tank 600 l, equipped with black polythene shreds as plant imitation
(Fig. 5), the second group in a 150 x 60 x 50 cm tank (450 l), and the last group in a
smaller 90 x 60 x 53 cm tank (300l). Conductivity and temperature were measured
daily.
2.3 Notopteridae
2.3.1 Notopterus notopterus
2.3.1.1 Specimens and breeding tanks
Some specimens of Notopterus notopterus (Fig. 6), originating from South-East Asia,
had already been present at the Humboldt University of Berlin, before the onset of these
investigations. These were nine females and five males with a total length of
198-269 mm and a total weight of 45.97–158.66 g. The additional 20 specimens
(10♀, 10♂) were purchased from a wholesale dealer (Aquarium Glaser in Rodgau, near
Frankfurt am Main). This group consisted of individuals ranging from 178–248 mm in
total length and 39.38–136.77 g in total weight.
Sexes of Notopterus notopterus can be distinguished by the shape of the genital papillae
(Weitkamp, 2005). The male’s narrow genital papilla is of reddish color and longer than
the rudimentary pelvic fin, while the female’s broader one is of whitish color and
shorter than its pelvic fin (Fig. 7). The fish were fed once a day on beef heart and/or
frozen chironomids.
Fig. 6. A female Notopterus notopterus with a total length of 23 cm. Scale bar = 2 cm.
Two tanks, one of 700 l and 400 l volume, respectively, were used to breed this species
with sex ratios 2:1 and 1:1 (female/male). Each tank had been disinfected before usage.
One breeding tank was equipped with black polythene shreds as plant imitation or
hiding places and some flat large stones as spawning substrates (Fig. 7a), whereas the
13
other tank was supplied with two large stones and sand, covering the entire bottom of
the tank (Fig. 7b).
Fig. 7. Adult male [a] and female [b] Notopterus notopterus; the genital papilla (arrowhead)
The acclimatization period lasted about six months. This period of time was used to
observe the selected specimens for breeding purposes and also to create the desired
breeding conditions. During this phase, the fish established individual territories.
Moreover, the male chose a female partner for courtship and mating.
2.3.1.2 Rearing
Eggs, embryos and larvae were attained from ten spawning occurrences. Directly after
each spawning, the eggs were removed from the breeding tank and transferred into
20 x 10 x 6 cm plastic jars covered with a plastic lid or into Petri dishes measuring
5-10 cm in diameter.
Fig. 8. Two experimental tanks of Notopterus notopterus. [a] A tank equipped with flat stones (x) and
black polythene shreds (arrow). [b] The tank was filled up with sand on the ground and equipped with
two large stones (x) as well as live plants and roots of Monstera.
They were afterwards placed in a thermostat (27 °C, no aeration system) until the larvae
started exogenous feeding. Larvae were fed for the first 7 days with fresh, newly
hatched Artemia nauplii. Subsequently on the eighth day of feeding, food supply was
substituted by older Artemia nauplii supplemented with small pieces of Tubifex. As the
14
larvae grew, they were transferred into a small tank (20 x 20 x 20 cm) and provided
with small pipes as hiding places. Once older than three months, the juvenile fish were
fed with small pieces of beef heart twice a week.
2.3.2 Xenomystus nigri
2.3.2.1 Specimens and breeding tanks
The first 16 (9♀, 7♂) fish arrived on 30 March 2009 and comprised a range in total
length of 85–120 mm (Fig. 9). The fish, originating from the Niger Basin, were
purchased from a whole sale dealer. This group was placed in a tank measuring
120 x 75 x 60 cm and one third of the water was regularly changed once a week. The
first tank was furnished with black polythene shreds as hiding substrates.
Two weeks upon arrival, some of the fish were infected with the ectoparasite
Ichthyiophthirius known as white spot disease. Medicinal treatment was performed for
three weeks, although three heavily infected fish died. The remaining fish recovered
well. Acclimatization was continued in a disinfected tank for about 60 days.
Fig. 9. Adult female Xenomystus nigri with a total length of 10.7 cm. White arrowhead points to the
swollen belly. Scale bar = 2 cm.
Fig. 10. The genital papillae in Xenomystus nigri can be used for the differentiation between [a] female
and [b] male; the male has a short protruding papilla (arrowhead) lying underneath the pelvic fin while
the female’s genital papilla is longer. Scale bar = 1 cm.
15
The second group was purchased on 6 May 2009 from a wholesale dealer
(Aquarium Glaser in Rodgau near Frankfurt am Main) consisting of 10 specimens
(9♀, 1♂) with a total length of 128–173 mm. The specimens were placed into a tank
measuring 90 x 60 x 53 cm, which was also supplied with black polythene shreds and
two PVC tubes as hiding places. Acclimatization period lasted about three months.
Xenomystus nigri has no sexual dimorphism, except for the genital papillae, which is
remarkably longer than the pelvic fin in females, while the male’s papilla is a short
protrusion lying underneath the pelvic fin (Nyonje, 2006) (Fig. 10). Fish were fed once
daily with live Chaoborus sp. and/or frozen chironomids. Selected specimens with
swollen bellies received several hormonal injections (see 2.8). Three tanks
(90 x 60 x 53 cm) were furnished with several PVC tubes, stones and woods. Each of
tanks comprised one male and female. The selected specimens were separated for two
weeks before the injection. Immediately after the injection, all specimens were returned
into their previous tanks.
2.4 Osteoglossidae
2.4.1 Osteoglossum bicirrhosum
2.4.1.1 Location and fishes
Study of the Silver Arowana (Osteoglossum bicirrhosum) (Fig. 11a) took place in
Florencia, Southeast Colombia for around 19 weeks. The samples were collected from
different ponds and/farms located at least 5 km outside of the city (El Doncello,
El Paujil , Montañita, Morelia, San Jose, San Vicente). Samples consisted of eggs and
juveniles of Osteoglossum bicirrhosum in different stages of development.
O. bicirrhosum can be kept together with different fish species such as
Oreochromis niloticus (family Cichlidae), Arapaima gigas (family Osteoglossidae),
Piaractus brachypomus
(family
Serrasalmidae),
Colossoma macropomum
(family Characidae),
Potamotrygon sp.
(family Potamotrygonidae)
and
Pseudoplatystoma fasciatum (family Pimelodidae). The fish were fed with pellets and
live guppies, ad libidum. Since tap water could not be utilized due to the high level of
Chlorine (CL), water taken directly from the river nearby was used. The water
temperature ranged from 26-31 °C depending on weather conditions and pH value
varied from 6.8–7.2. About one meter above the surface the ponds were covered with
nets for protection from predators.
2.4.1.2 Collection of samples
Prior to collecting the samples, some observations of fish behaviour had to take place.
Guarding males ignored the pellets, as they cannot swallow them. Every specimen of
16
Silver Arowana with a closed mouth and swollen hard jaw contained eggs, embryos, or
even free embryos. The volume of the ponds (Fig. 11b) ranged from 600 to 3000 cbm.
Collecting the samples always took place either in the early morning or late afternoon in
order to avoid the strong sun.
The heavy rainy season is used as a signal for the start of the spawning season of
Osteoglossum ssp by the farmers. Using a long net (ca. 45 m long) the fish were driven
to the edge of the pond (Fig. 11c). The selected specimen was gently touched on the
bottom of the jaw to stimulate the opening of its mouth. The eggs or embryos found in
its mouth were cautiously poured into a plastic jar filled with water from the pond
(Fig. 11d) and shielded from direct sunlight. Subsequently they were carefully put into
a transparent plastic sac filled with clean water of the breeding pond and supplied with
oxygen (Fig. 11e). Eggs and embryos were kept separately.
Fig. 11. Collecting samples on the farm. [a] Adult specimen of Osteoglossum bicirrhosum with a total
length of 73 cm and 4 years old, located in Paujil. [b] This pond contained 40 Silver Arowana and was
covered with a net for protection from predators. [c] By using a long net pulled by hand, all fish were
gathered to the edge of the pond. [d] Transferring the eggs into a plastic jar. [e] Collected eggs were put
into a transparent plastic bag ready to be transported to the laboratory.
17
They were directly transported to the city for further investigation. The transport itself
took around 2–3 hours. Unfertilized eggs or dead specimens (recognized by white
colouring and gradually damaging process of the egg envelope) were immediately
removed from the remaining specimens (Fig. 12).
2.4.1.3 Rearing and documentation
To raise the eggs (Fig. 13a) and embryos (Fig. 13b), the samples were respectively put
into two separate aquaria (25 x 15 x 20 cm) (Fig. 13). Each aquarium was equipped
with a filter, heater, thermometer and a lid made from Styrofoam. Especially the freshly
captured eggs were put into a small transparent glass container inside the aquarium that
was equipped with two filter devices: one to produce the oxygen and the other, which
was connected with a tiny tube, to produce a slight current for maintaining a continuous
movement of the eggs (Fig. 13a, b).
Fig. 12. Separation of samples in Osteoglossum bicirrhosum. [a] Freshly collected eggs and [b] embryos
were directly separated from the unfertilized eggs which can be recognized by their white colour
(arrows); these pictures were taken from two different farms.
Fig. 13. Aquarium set up containing [a] freshly collected eggs and [b] embryos of
Osteoglossum bicirrhosum. Transparent glass container (x), heater (arrow), thermometer (*), two filter
devices (arrowhead) and a Styrofoam lid (**) were used to maintain stable conditions for raising the eggs
and embryos.
18
Water used for the aquaria, taken from the local farm, was regularly changed to uphold
its quality. The water temperature in the aquarium was kept constant at ca. 28 °C with
pH ranging from 6.8–7.2; water level varied from 15–18 cm. Exogenous feeding
commenced earlier than expected even though the yolk sac had not been resorbed yet.
During this period the embryos were fed three times a week with flakes. Once the yolk
sac had been completely resorbed, the juveniles kept on being fed twice a day with
flakes and/or substituted with small guppies, ad libidum.
On 22 February 2011, 16 dead specimens were collected from a nearby pond consisting
of two specimens of Osteoglossum bicirrhosum and 14 specimens of
Osteoglossum ferreirai. They were measured and dissected two hours after having been
taken out of the pond. The time delay was due to the transport from the farm to the
laboratory. In vivo observations on freshly captured eggs and embryos were
photographed by using a stereomicroscope Nikon SMZ 7454 connected with
Optika Vision Pro software. Meanwhile photos of the development of embryos, juvenile
and adult were captured by using Sony TX7.
2.5 Variation and measurement of environmental parameters
Variation of environmental parameters to induce the gonad maturation was stimulated
with the method developed by Schugardt and Kirschbaum (2004), namely through
manipulation of the conductivity of the water. Continuous decreasing conductivity was
achieved by adding demineralised water to the water in the experimental tanks. The
water level (WL) was kept constant through an overflow system. Conductivity (C) and
temperature (T) were measured daily by using a conductivity meter
(WTW Tetracon 325); photoperiod was set at 12L:12D.
2.6 Behaviours
Observation of social, courtship and spawning behaviors was performed during day
time. If courtship was intensely seen, evening and night observations were necessary.
2.7 Maturity coefficient (MC)
The gonad’s maturity coefficient (MC) of dissected-specimens measured in percentage:
(gonad weight/ total weight-total gonad weight) x 100, was used to verify and evaluate
the effect of environmental factors on gonad development.
19
2.8 Hormonal treatment
Some trials of hormonal treatment were applied during the experimental period to a
number of specimens of Xenomystus nigri. The selected specimens had swollen bellies
and showed courtship behaviour regularly for several weeks. They were isolated into a
tank of 90 x 60 x 53 cm. In order to obtain ovulated eggs and mature sperm, the selected
females and males were injected one after another by using ovaprim
(Vancouver, Canada), a GnRH analogue combined with domperidone (a dopamine
antagonist). Mature looking females and males (with large abdomen) were anaesthetised
with ethylenegylcol monophenyl ether. The anaesthesia was started with the males. Two
doses of the GnrRH hormone were injected intramuscularly at 50µg/kg for female with
8–10 g in body weight. Three units of hormone were injected at 75 µg/kg for male with
body weight of 10 g. Total weight and total length of each specimen were documented
and injection was applied on the left side of the body into the dorsal musculature. After
hormone injections the females and males were returned into the same tanks. Hand
stripping was applied to the same specimens, if no spawning occurred on the following
day. Its abdomen was kept dry and hand stripped applied gently on the area of abdomen.
2.9 General photography
All fertilized eggs, development stages of the embryo and larval development were
made with two different cameras: a Leica S6E binocular fitted with a Canon PC1048
micro-camera and a Canon Powershot S50, digital camera mounted on a Leica L2
Stereomicroscope. All pictures taken from the laboratory showing the representative
juvenile and adult fish as well as their tanks and in situ condition were photographed by
using a Canon EOS 350D digital Camera. Photographs were mostly taken of
anesthetised specimens.
2.10 Terminology
The terminology of early ontogeny which was applied here is based on the concept of
Balon (1975).
20
3 RESULTS
3.1 Family Mormyridae
3.1.1 Petrocephalus soudanensis
The breeding experiment with Petrocephalus soudanensis lasted 371 days (Fig. 14).
Conductivity was decreased three times throughout the breeding experiment with the
first period starting from 804 (day 1) to 114 µS/cm (day 80), imitating the rainy season
to provoke gonad maturation. The conductivity was gradually increased up to
700 µS/cm (day 117). The second period was started on day 118 and lasted until day
175 with minimum conductivity of 109 µS/cm. The conductivity was increased again
steadily for a longer period of time from day 176 to day 294 at 725 µS/cm. The last
experimental period started on day 295 and within 40 days there was significantly
decreased conductivity to 89 µS/cm, which was followed by a drastically increasing
conductivity from 141 (day 351) to 750 µS/cm (day 353) then again slowly continued
increasing to 788 µS/cm. Water level was kept constant at 41 cm and the temperature
varied from 24.5 to 27.5 °C.
Fig. 14. Course of environmental factors, decreasing conductivity (C), constant water level (WL), and
temperature (T) in the breeding experiment with 1 female and 3 males of Petrocephalus soudanensis to
provoke gonadal recrudescence and spawning. Around day 18 the female showed a swollen belly as
indication for gonad maturation. Courtship behaviour was seen, however spawning was not.
21
Around day 18, the female showed for the first time a swollen abdomen indicating
gonad maturation. This followed by intense courtship behaviour five days after (day 23)
and lasted for 53 days; a pair was actively seen together during day and night time.
Courtship behaviour discontinued during the increasing conductivity. The second
swollen belly appeared on day 126 after the onset of the second rainy season and
followed by courtship behaviour on day 127 to day 173. On day 183 the female showed
a regressed abdomen concomitantly by increasing conductivity. Later on, the belly was
seen starting to swell up on day 316 and continuing to do so until day 331, on which a
really huge belly was seen, but, unlike the previous symptom, was not followed by
courtship behaviour this time. However, even though some intense swollen bellies and
courtship behaviour occurred (Fig. 15, 16), still no spawning took place.
Fig. 15. Courtship behaviour seen in Petrocephalus soudanensis during the breeding experiment. [a] A
pair was found daily inside a PVC tube on the left side of the tank. [b] They swam and nudged together
through the tube. Some actions were captured in the afternoon during courtship behaviour; [c-d] the pair
was circling around and swimming alongside for a while on the right side of the tank before returning to
the isolated plastic tube on the left side of the tank. The swollen abdomen of the female is visible.
All observed courtship was seen in the morning or prior midday and in the afternoon.
The female stayed with one male during the whole courtship period. They were found
inside of a PVC tube on the left side of the tank (Fig. 15a). They swam and nudged
together through the tube (Fig. 15b). Once the female left the tube and swam to the
right side of the tank, the male directly followed the female, either from behind or from
22
the other way around. They were circling around and swimming alongside for a while in
this area (Fig. 15c-d), and then they returned again to the same PVC tube. The pair
established a territorial behaviour where they mostly occupied the bottom area around
the plastic tubes. The two other males were not involved in courtship behaviour.
Fig. 16. Illustration of the swimming movements (arrows) during social and courtship behaviour of
Petrocephalus soudanensis (1♀, 3♂); [a] Front view; [b] upper view.
After almost a year of experimenting, the breeding experiment was stopped since the
female showed a thinner belly and spawning had not occurred. Some time after the
onset of the imitation of the dry season (increase of conductivity) courtship behaviour
also decreased. The swollen belly and the courtship behaviour indeed indicated
successful gonad maturation. Spawning, however did not take place.
3.1.2 Petrocephalus catostoma
The first breeding experiment lasted 257 days comprising one female and four males.
The course of the environmental factors is shown in Fig. 17. Conductivity was
decreased from 875–155 µS/cm with pH value varying from 8.4–7.8, whereas
temperature fluctuated around 24.1–27.8 °C. Water level was kept constant at 40 cm. In
the course of the breeding experiment one male and the female have established
territories next to each other. Courtship behaviour as in P. soudanensis was not found.
23
Fig. 17. Course of environmental factors, decreasing conductivity (C), constant water level (WL), and
temperature (T) in the breeding experiment group I with a female and four males of
Petrocephalus catostoma to provoke gonadal maturation and spawning. The female showed a swollen
belly as first indication of gonad maturation on day 66. Neither courtship behaviour nor spawning
occurred during the breeding experiment.
Fig. 18. Illustration of the position of the territories of 1 female and 4 males of Petrocephalus catostoma
during the day time, [a] Front view; [b] upper view.
24
The female’s belly did swell up during the 66 day experiment. The territories of the five
fish are shown in Fig. 18. However, there was only a change in the position of the
territories of 1 male and the female during the course of the breeding experiment.
Within seven months, the specimens did show declining growth in terms of total length
and total weight (see Table 1).
Fig. 19. Course of environmental factors, decreasing conductivity (C), constant water level (WL), and
temperature (T) in the breeding experiment group II with a female and two males
Petrocephalus catostoma to provoke gonadal maturation and spawning. Neither courtship behaviour nor
spawning occurred during the breeding experiment.
Table 1. Measurement of total length and total weight of Petrocephalus catostoma during the first
breeding experiment
No.
1
2
3
4
5
Sex
♂1
♀
♂2
♂3
♂4
20.03.2009
TW (g)
20.13
22.89
23.32
18.41
21.23
TL (cm)
11.60
12.10
11.80
11.10
11.70
Sex
♂1
♀
♂2
♂3
♂4
16.06.2009
TW (g) TL (cm)
18.72
11.60
21.43
12.10
20.58
11.80
16.91
11.10
19.92
11.70
Sex
♂1
♀
♂2
♂3
♂4
12.10.2009
TW (g) TL (cm)
16.29
11.60
20.71
12.10
20.48
11.90
14.45
11.10
16.49
11.70
The second breeding experiment of P. catostoma lasted for 223 days involving the
female and two males (Fig. 19), which were already used for the first breeding
experiment (Fig. 18). The conductivity varied from 779 to 140 µS/cm, pH value from
25
8.3-6.8 and the temperature from 24.1–30 °C. Similar to the first breeding experiment,
no unusual behaviour was observed throughout the entire period. Indication for the
female’s swollen belly was observed here on day 17, way earlier than in the first
breeding experiment. However, neither courtship behaviour nor spawning was observed
afterwards.
3.2 Family Pantodontidae, Pantodon buchholzi
Figure 20 and 21 present the course of applied environmental factors; water level (WL),
conductivity (C) and temperature (T) in breeding group I (7♂, 5♀) during period of 328
days and in breeding group II (1♂, 2♀) within 221 days of breeding experiment,
respectively. Two fish (1♂, 1♀) of the first group showed signs of courtship behaviour
during the decreasing conductivity. However, intense courtship lasted only for several
days starting from day 54 and lasting until day 67. This pair was often seen swimming
close to each other and the male swam actively to attract the female’s attention. This
observation took place mostly during daytime.
The two experimental tanks had been equipped differently, which provoked two
different types of social behaviours. In the first group of P. buchholzi, the tank was
furnished with black plastic shreds and several plastic tubes. The fish adapted very well
to this environment, hiding between the plastic shreds. During feeding, the fish reacted
differently compared to the second group, whose tank had not been supplied with
decoration. These fish calmly waited for the crickets, which were running on the water
surface, to approach them. On the other hand, the fish of the second group swam
actively and very quickly to catch the crickets on the water surface.
In addition, three breeding experiments with combination (C) of I (1♀, 1♂),
II (5♀, 4♂), and III (4♀, 5♂) were performed in three different tanks of 90 x 60 x 53 cm
for a time period of two months with several trials of drastically decreasing
conductivity. However, neither a swollen belly nor courtship behavior was observed
during these trials. Six (2♀, 4♂) specimens were kept in the tank for nearly eight
months, whereas the rest of specimens were put to death and dissected (see Table 2).
The six specimens were measured monthly in total length and total weight. The fish
often had been seen swimming on the surface and somehow lost their appetite towards
the crickets. However, neither courtship nor spawning took place. Four specimens died
one after another due to some unidentified illnesses.
26
Fig. 20. Course of environmental factors, decreasing conductivity (C), constant water level (WL), and
temperature (T) in the breeding experiment group I (7♂, 5♀) of Pantodon buchholzi to provoke gonadal
maturation and spawning. No spawning occurred during the breeding experiment.
Fig. 21. Course of environmental factors, decreasing conductivity (C), constant water level (WL), and
temperature (T) in the breeding experiment group II of Pantodon buchholzi with two females and a male,
to provoke gonadal maturation and spawning. No spawning observed.
27
Fig. 22. In situ condition of the gonads in Pantodon buchholzi. [a] Light greenish female gonads
surrounded with fat tissue, total length 10.3cm whereas [b] some gonads were found of whitish color. [c]
Gonads taken out left (0.22 gr) and right (0.21 gr). [d] Testes freshly taken out, left (0.23 gr) and right
(0.30 gr). Scale bar = 20 mm (a), 10 mm (b, c, d).
In situ situation in all specimens of Pantodon buchholzi demonstrated two different
colours of the female’s gonad - greenish and whitish (Fig. 22). The ovary is made up of
left and right parts. The total gonad weight of the female (TGW) ranges between
0.08-0.73 g (left) and between 0.1–0.95 g (right), whereas the male’s total gonad weight
(TGW) varies between 0.02–0.23 g (left) and 0.02–0.3 g (right). Table 2 below presents
the percentage of the maturity coefficient (MC) of 41 dissected specimens including
total length (TL), total weight (TW) and total gonad weight (TGW) of
Pantodon buchholzi.
28
Table 2. Maturity coefficient (MC) of dissected Pantodon buchholzi at the end of the experimental
period, with total length (TL), total weight (TW), and total gonad weight (TGW) data from all groups (I,
II, CI-Combination I, CII-Combination II, CIII-Combination III, R-Rest)
No
Sex
TL (cm)
TW (cm)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
♂
♂
♂
♂
♂
♂
♂
♀
♀
♀
♀
♀
♂
♀
♀
♂
♀
♂
♂
♂
♂
♀
♀
♀
♀
♀
♂
♂
♂
♂
♂
♀
♀
♀
♀
♀
♀
♂
♂
♂
♂
10.00
9.90
9.80
10.00
10.00
9.70
9.10
9.30
9.00
9.50
9.30
9.00
9.30
9.30
9.00
10.00
8.00
10.00
8.80
9.30
9.60
8.50
8.70
8.50
10.30
10.00
9.50
8.00
9.40
9.00
9.00
9.30
9.60
9.80
11.00
9.80
9.50
10.30
10.30
10.50
10.90
6.62
7.41
6.42
6.81
7.53
6.54
5.49
6.71
6.01
5.40
5.43
5.80
6.55
4.58
5.80
5.79
3.96
7.35
5.62
6.24
6.60
4.22
5.37
6.07
7.68
8.90
6.18
4.42
6.34
4.34
4.40
6.00
7.76
7.48
9.36
6.64
6.30
7.68
7.93
9.77
9.78
TGW (g)
L
0.07
0.09
0.06
0.09
0.05
0.08
0.15
0.43
0.31
0.13
0.09
0.06
0.08
0.13
0.12
0.02
0.08
0.08
0.09
0.09
0.14
0.21
0.25
0.42
0.22
0.73
0.07
0.05
0.08
0.13
0.10
0.11
0.60
0.29
0.22
0.31
0.20
0.22
0.11
0.23
0.18
TW (cm)
R
0.09
0.12
0.11
0.08
0.10
0.11
0.12
0.53
0.36
0.09
0.10
0.10
0.10
0.19
0.13
0.02
0.11
0.11
0.10
0.07
0.18
0.27
0.27
0.55
0.21
0.95
0.11
0.05
0.09
0.14
0.10
0.10
0.70
0.50
0.38
0.40
0.30
0.21
0.16
0.30
0.27
MC (%)
L
R
1.10
1.38
1.23
1.64
0.94
1.70
1.30
1.18
0.67
1.30
1.30
1.70
2.80
2.23
6.80
8.60
5.40
6.37
2.70
1.70
1.70
1.90
1.05
1.75
1.20
1.70
2.90
4.32
2.10
0.20
1.34
1.34
2.06
2.85
1.20
1.50
1.60
1.26
1.50
1.60
2.16
2.80
5.23
6.80
4.80
5.30
7.40
9.90
2.90
2.80
0.80
1.20
1.13
1.79
1.14
1.14
1.27
1.40
3.30
3.33
2.30
2.30
1.80
2.14
7.80
9.10
4.00
7.10
2.40
4.20
4.90
6.40
3.30
5.00
2.90
2.80
1.40
2.06
2.40
3.16
1.87
2.83
Group
I
I
I
I
I
I
I
I
I
I
I
I
II
II
II
CI
CI
C II
C II
C II
C II
C II
C II
C II
C II
C II
C III
C III
C III
C III
C III
C III
C III
C III
C III
R
R
R
R
R
R
29
3.3 Family Notopteridae
3.3.1 Xenomystus nigri
The first breeding experiment lasted for 470 days. Decreasing conductivity took place
for three times within the first period (Fig. 23) ranging from 901 to 301 µS/cm, with
temperature varying from 24.3–29.9 °C and pH value ranging from 8.4–6.6. Water level
was always kept constant at 40 cm.
Fig. 23. Course of environmental factors, decreasing conductivity (C), constant water level (WL), and
temperature (T) in the breeding experiment group I for a 16-month period of Xenomystus nigri (9♀, 7♂),
to provoke gonadal maturation and spawning. First swollen belly was seen in two females on day 36,
arrow.
First indications for the swelling bellies of three females was seen on day 36 at
342 µS/cm. Courtship behavior took place for a few times during the experiment,
however, spawning was never observed although some females and males continued
showing obviously swollen bellies. Three females died throughout the course of the
experiment.
30
Fig. 24. Course of environmental factors, decreasing conductivity (C), constant water level (WL), and
temperature (T) in the breeding experiment group II for 13 months period of Xenomystus nigri (9♀, 1♂),
to provoke gonadal maturation and spawning. First three females with swollen bellies were seen on day
70, arrow.
The three selected pairs, showing significantly swollen bellies, were subjected to the
application of hormonal injection. They were pair I (♂1: TW= 26.7 g; TL= 16.5 cm and
♀1: TW= 29.14 g; TL= 17 cm), pair II (♂2: TW= 21.58 g; TL= 15.5 cm and ♀2:
TW= 29.14 g; TL= 17 cm), and pair III (♂3: TW= 21.52 g; TL= 15.5 cm and ♀3:
TW= 24.67 g; TL= 16.9 cm). Five hormonal injections were given to these three pairs
on day 389, 395, 402, 408, and day 415. Apparently those five hormonal injections
seemed to have no particular impact on inducing spawning, although intense courtship
behaviours were often observed within these three pairs. The pairs indeed were seen
swimming very closely to each other during day time. In fact, double of the
concentration of the regular hormonal injection was given on the last day (day 415) of
the trial to all of the selected specimens, however still nothing happened. Nevertheless,
triggered by decreasing conductivity, gonad maturation could successfully be induced
on this species, which was apparent by its significant changes around the bellies.
The second breeding experiment lasts for 394 days consisting nine females and a male.
The first swollen belly was observed at day 70 in three females. Similar to the breeding
experiment 1, no spawning occurred. Two females died throughout the course of the
experiment.
31
Fig. 25. In situ condition in Xenomystus nigri. [a] Female’s gonad occupies almost 80 % of internal
organ, total length 16 cm and total weight of 22.38 g. [b] Gonad consists of different stadium of oocytes,
larger magnification. [c] Male’s in situ with undeveloped testes, total length 15.4 cm and total weight of
17.27 g. Scale bar = 20 mm (a), 10 mm (b, c, d).
Figure 25 shows the in situ condition of Xenomystus nigri presenting the ovary
(Fig. 25a) and testis (Fig. 25b). X. nigri possesses only a single ovary positioned on the
left side of the abdominal cavity. The ovary consisted of different stadia of oocytes. A
single testis was found laterally in a similar position in the male's abdomen, lying on the
apex of the coiled digestive tract. Maturity coefficient value of figured female is 7.7 %
and the male has 0.23 %. Table 3 presents the percentage of the maturity coefficient
(MC), including total length (TL), total weight (TW) and total gonad weight (TGW), of
21 dissected specimens of Xenomystus nigri at the end of all experiments.
32
Table 3. Maturity coefficient (MC) of dissected Xenomystus nigri at the end of experimental period, with
total length (TL), total weight (TW), and total gonad weight (TGW) data from breeding group I and II
No
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
Sex
♀
♀
♀
♀
♀
♀
♂
♂
♂
♂
♂
♂
♂
♀
♀
♀
♀
♀
♀
♀
♂
TL (cm)
14.30
13.90
15.70
12.80
16.30
17.50
14.90
14.00
13.70
14.60
17.00
15.50
15.50
16.40
15.40
14.80
15.30
17.80
16.90
14.60
15.70
TW (g)
15.80
14.31
22.00
10.60
22.38
28.28
17.78
14.61
13.60
16.54
27.24
8.62
17.27
21.42
18.45
15.58
22.05
19.25
24.11
15.82
21.84
TGW (g)
0.94
0.44
1.56
0.50
1.60
1.06
0.20
0.03
0.02
0.10
0.10
0.10
0.04
0.11
0.11
0.06
1.70
0.68
0.66
0.78
0.30
MC (%)
6.50
3.20
7.63
4.95
7.70
3.89
1.10
0.20
0.15
0.30
0.40
0.80
0.20
0.50
0.60
0.40
8.30
3.66
2.80
5.18
1.40
Breeding Group
I
I
I
I
I
I
I
I
I
I
I
I
I
II
II
II
II
II
II
II
II
3.3.2 Notopterus notopterus
3.3.2.1 Reproduction
In N. notopterus only the left gonad is developed (dissection of 16 specimens (9♀, 7♂)).
The first breeding experiment was performed with one male and two females (Fig. 26).
The variation of conductivity was fulfilled, based on experiments with mormyrids and
gymnotiforms (Kirschbaum and Schugardt, 2004), which had shown that decreasing
conductivity can elicit gonad maturation. A swollen belly as a sign for gonad maturation
was seen for the first time in female No. 1, 33 days after the onset of the experiment.
For observational purposes, a tiny part of the caudal fin of this female was cut out to
easily identify the specimen. First time spawning occurred on day 198 of the
experimental period, followed by 19 spawning events with irregular intervals (see
Table 4) within a five-month period. Female No. 2 spent most of the time alone on the
other part of the tank without interrupting the pair during courtship and spawning
behaviour.
The three fish were 269 mm (♀ No.1), 236 mm (♀ No. 2), and 250 mm (♂) in total
length (TL) and varied in total weight of 99.25 g, 83 g and 118 g, respectively. On the
33
following day the conductivity was decreased from 780–620 µS/cm (Fig. 14) to start the
experiment.
Fig. 26. Course of environmental factors conductivity (C), water level (WL), and temperature (T) during
339 days in breeding tank I containing two females and one male of Notopterus notopterus. 20 spawning
occurrences of the female No. 1 were observed within a-5-month period. The breeding experiment was
commenced after a 3 months acclimatization period. Note the first swollen belly of the female No. 1
(Arrow). * refers to observed spawning.
The second breeding experiment comprised one male (TL= 261 mm, TW = 110 g) and
female (TL= 236, TW= 83 g) (Fig. 27). A swollen belly of the female concurrently with
courtship behaviour was first observed on day 62. However, no spawning occurred
during this initial courtship period. Around 15 times, intense courtship behaviour was
observed mostly from 12.00–6.00 pm throughout a five-month period. However,
spawning occurred only six times. The first spawning of this pair was found on day 178,
around 5.30–7.00 pm, whereas the following spawning mostly took place from 8.00 to
2.30 pm. Maturity coefficient (MC) values of specimens used in breeding experiment I
at the end of the experimental period are 8.46 % (♀1), 8.75 (♀2) and 1.12 (♂),
corresponding to Figure 26. Whereas in breeding experiment II (Figure 27) the
maturity coefficient (MC) for the single male and female are 0.76 % and 7.52 %,
respectively.
34
Fig. 27. Course of environmental factors during 331 days, conductivity (C), water level (WL) and
temperature (T) in breeding tank II containing one female and one male Notopterus notopterus. An
irregular spawning interval was observed for 6 times within a 6-month experimental period. The breeding
experiment was started after 40 days of acclimatization. Note the first swollen belly of the female
(Arrow).* refers to observed spawning.
Courtship behaviour was regularly observed throughout the entire daytime. Overall
there were five steps of courtship behaviour observed during these breeding
experiments: 1) The male followed or swam alongside the female as the female often
swam faster than the male, 2) Male touched female’s belly with his mouth several times
while the female actively swam up and down in front of the male, 3) Male quivered the
female by swinging rapidly its tail against the side of the female’s body, to which the
female reacted by swimming quickly to the other side of the tank and then returning
directly to its previous position, 4) A male and a female stayed still in a corner of the
tank until the male suddenly swam to the other side on its own, 5) The male later on was
actively approached by the female and led the female to the spawning site. Two
representative pictures of courtship behaviour from the second breeding experiment are
presented in Figure 28. In the particular case of the second breeding pair, mostly during
this intense courtship behaviour the male was actively seen preparing a suitable
spawning site, by removing gravels and cleaning the spawning site with its mouth
(Fig. 29).
35
Fig. 28. Two elements of courtship behaviour in Notopterus notopterus. [a] Around the spawning site the
male touched the female’s abdomen with its mouth and [b] the pair remained together on the right side of
the tank.
Fig. 29. Intense removing gravels activity was found in breeding experiment II. Male was actively seen
collecting gravel the whole day and removed it to somewhere else nearby. These activities [a, b] will
stop, once the preferred ground is completely gravel-free.
The 20 successful spawning events of the first breeding experiment were observed
during day time. The pair seemed ready to spawn as the female showed a swollen belly
and the male was apparently attracted by the female. Soon after the female had laid
some eggs on the preferred substrate, the male quickly fertilized the eggs while the
female was still nearby. When spawning was finished, the male remained at the
spawning site, while the female left for the other side of the tank. The genital papilla of
the female appears bigger than usual during courtship and spawning. It is approximately
5–7 mm in length.
36
The complete sequence of the spawning number two (11.01.2010) from the first
breeding pair is illustrated in six elements as shown in Figure 30: a) Male pushed the
female with his mouth to the spawning substrate, b) after the male fertilized the eggs,
the pair was seen together very close to their newly spawned eggs, c) the female laid
some eggs for the second time, which were then directly fertilized by the male, d) male
pushed the female with his head to lay some eggs on the other side of the rock, the
female seemed to be in a leaning position of 30° angle, e) after the female laid some
eggs, the male directly fertilized the eggs, while the female still stayed very close,
somehow touching the male’s belly with its head, f) the female laid eggs for the last
time in this sequence.
Table 4. Overview of spawning (Sp) events in breeding group I (2♀, 1♂) and breeding group II (1♀, 1♂)
with number of eggs, pH value and temperature (T) per individual spawning
Breeding Group I
Sp. Nr.
Sp. Date
1
04.01.2010
50
2
11.01.2010
3
Breeding Group II
Nr. Eggs pH
T (°C)
Sp. Nr.
Sp. Date
Nr. Eggs
pH
T (°C)
8.5
26
1
16.12.2009
87
6.5
28
68
8.2
26.2
2
23.12.2009
75
7.0
28
19.01.2010
90
7.9
25.7
3
14.01.2010
50
8.2
27.8
4
21.01.2010
50
7.7
25.3
4
17.02.2010
47
7.8
27.8
5
02.02.2010
50
7.7
27.8
5
28.02.2010
45
7.6
27
6
03.02.2010
25
7.7
27.1
6
27.03.2010
51
6.0
27
7
16.02.2010
74
7.5
26.2
8
17.02.2010
30
7.5
26.3
9
25.02.2010
105
7.0
25.9
10
02.03.2010
225
7.0
27
11
12.03.2010
113
6.5
24.9
12
17.03.2010
27
6.5
26.4
13
23.03.2010
120
6.7
26.2
14
28.03.2010
40
7.3
26
15
29.03.2010
180
7.3
25.6
16
04.04.2010
70
7.6
26.1
17
13.04.2010
15
7.9
26
18
10.05.2010
118
8.3
26.3
19
17.05.2010
30
8.0
25.7
20
25.05.2010
80
1560
7.7
26.3
355
Regarding most of the spawning events of the first breeding pair, nearly all the eggs
were spawned at the same substrate and location. The eggs were attached to the bottom
side of larger stones. Nevertheless, in the last two spawning events of this breeding pair,
eggs were found outside of the common area: on the edge of the filter (Fig. 31c), even
though some larger stones were positioned nearby. The female deposited circa 15–225
eggs per spawning and 1560 eggs during the 20 spawning occurrences (see Table 4).
37
Meanwhile, the second breeding pair always spawned underneath or nearby large
stones. Within six spawning events, the female deposited around 355 eggs in total.
Detail numbers of each spawning is presented in Table 4.
Parental care is performed by the male. This implies defending the nest against other
specimen in the tank and guarding the eggs (Fig. 32). For two days after spawning the
male feeds on significantly less food and protects the eggs aggressively, also against the
female.
Fig. 30. Spawning sequence of Notopterus notopterus. Egg deposition occurred in different positions,
while the male accompanied the female in every action. [a] The male gently pushed the female on its
belly with its mouth, to lead her to the desired spawning substrate. [b] Both were seen staying very
closely together after the male fertilized the eggs. [c] The second spawning of the female, followed by the
fertilization of the male. [d] Male pushes the female again to other substrate nearby the first location of
spawning. [e] The male directly fertilized the eggs. [f] The female spawns the eggs for the last time on
this occasion.
38
3.3.2.2 In situ condition of mature and immature (F1) specimens
Figure 33 is the in situ condition of a dissected female Notopterus notopterus
presenting an immature gonad (Fig. 33a) and a mature gonad (Fig. 33b), respectively.
N. notopterus possesses only one single ovary positioned laterally in the abdominal
cavity on the coiled intestine on the left side of the fish. The immature gonad contains
mostly oocytes in stadium 1 with their very dominant whitish colour as seen in
Figure 33a. The picture of the immature gonad was taken from a female of 12 cm in
total length and total weight of 10.32 g.
Fig. 31. Preferred spawning sites of Notopterus notopterus. [a] The eggs attached to the underside
(double arrowhead) [b] on the edge of a large stone. [c] Eggs were also found on the edge of the filter.
Scale bar = 20 mm (a); 4 mm (b); 10 mm (c).
Fig. 32. Parental care of Notopterus notopterus performed by the male. [a] Male in breeding tank 1 and
[b] Male in breeding tank 2. Both males were always seen nearby the spawning substrates, note the
male’s genital papilla (white arrow) and newly spawned eggs (black arrowhead).
39
Fig. 33. In situ condition in female Notopterus notopterus. [a] An immature gonad, containing only
oocytes in stadium 1, total length (TL) 12 cm. [b] Mature gonad consists of different stadia of oocytes,
TL= 23 cm. Black arrowhead points the gonads. Scale bar = 20 mm (a), 10 mm (b, c, d).
The mature gonad of a female of 23 cm total length and 111.5 g weight was yellowishorange, occupied almost all of the space in the abdominal cavity, showed oocytes in
different stages and was surrounded with fat tissue. Maturity coefficient values for the
two females were 0.68 % and 8.46 %, respectively. Table 5 presents the percentage of
the maturity coefficient (MC) in 11 dissected specimens of Notopterus notopterus.
Table 5. Maturity coefficient (%) of dissected Notopterus notopterus, as relating to total length (TL),
total weight (TW), and total gonad weight (TGW) data
No
1
2
3
4
5
6
7
8
9
10
11
Sex
♂
♂
♂
♂
♂
♀
♀
♀
♀
♀
♀
TL (cm)
23.70
23.70
27.20
26.00
25.50
24.20
25.20
24.50
21.60
22.80
22.10
TW (g)
129.94
115.44
170.40
156.84
161.64
124.37
131.47
133.80
82.15
108.39
112.40
TGW (g)
0.45
0.77
1.26
0.83
1.18
5.68
7.44
6.54
2.76
5.63
7.64
MC (%)
0.34
0.67
0.74
0.53
0.73
4.80
5.90
5.13
3.47
5.47
7.29
3.3.2.3 External features of the egg
Fertilized eggs (Fig. 34) are adhesive, yellowish, and spherical with 3.8–4 mm in
diameter. The egg envelope has many external ridges which are centred around the
micropyle (Fig. 35).
40
Fig. 34. Newly spawned eggs of Notopterus notopterus. [a] The egg envelope has many spiralling ridges
originating from the micropyle. [b] The micropyle (m), located at the animal pole, is clearly marked as an
opening in the egg envelope of the ovulated egg. [c] View of an egg close to the animal pole. Arrowhead
points to the micropyle. Scale bar = 1 mm.
Fig. 35. Micropyle of Notopterus notopterus. [a] Larger magnification of Fig. 34b, note the depth of the
micropyle (m) (arrowhead). [b] Larger magnification of Fig. 34c, showing the spiralling ridges seen from
above. ev= egg envelope. Scale bar = 1 mm.
3.3.2.4 Development
Eggs were incubated at 27 °C. The terminology of the development of
Notopterus notopterus follows the classifications of Balon (1975). There are five
periods: the embryonic period (I), the larval period (II), the juvenile period (III), the
adult period (IV) and the senescent period (V). The last period will not be discussed in
this work due to the lack of information on this period in N. notopterus. Time of
spawning is used as the time zero in age determination; it is presented as hrs:min or
days.
3.3.2.4.1 The embryonic period
3.3.2.4.1.1 The cleavage phase
Stage 1: zygote-one cell
At 1:10, a discrete and distinct brownish pattern emerged to the top of the egg, the
blastodisc as characteristic of bipolar differentiation. Beneath the blastodisc, the
cytoplasm strictly joined in the yolk due to the formation of the prospective yolk
syncytial layer (Fig. 36a).
41
Stage 2: two-blastomere
At 2:00, the earliest cleavage furrow divides the blastodisc into two identical
blastomeres, the two-cell stage (Fig. 36b).
Stage 3: four-blastomere
The next cleavage furrow yields four equivalent blastomeres and is situated in a right
angle to the first cleavage furrow (Fig. 36c, 2:20).
Stage 4: eight-blastomere
The third cleavage occurs at 2:42 and produces eight blastomeres, arranged in two
parallel rows of four cells each (Fig. 6d).
Stage 5: early morula–16 blastomeres
The furrows of the fourth cleavage are oriented roughly horizontally to oblique
(Fig. 36e). The germ consists of 16 blastomeres at 3:45.
Stage 6: late morula
The fifth cleavage occurs at 4:30. The following cleavage furrows developed into
asynchronous producing unequal blastomeres in size and the germ comprises up to circa
32 blastomeres (Fig. 36f).
Stage 7: blastula
In late specimens of this stage, at 6:25, the blastodisc shows a definite pebbled
emergence and its upper exterior side slightly forms a dome-shape. It is indeed
distinguished from the yolk cell by a ring-like demonstrating the formation of the yolk
syncytial layer (Fig. 36g).
Stage 8: flat blastula
The blastodisc divides into two types of cells at 7:40. The primary cell population
recognized is the enveloping layer. It mostly originates from the superficial cells of the
blastoderm that form an epithelial sheet with a mono-cellular layer. The deep layer of
cells is located beneath the internal surface of the enveloping layer (Fig. 36h). Shortly
after, the margin of the blastoderm expands externally and the annular groove at the
blastoderm-yolk junction vanishes. This modification is the most noticeable sign
indicating the start of the epiboly.
42
Stage 9: late blastula
At this stage the deep cells have multiplied producing a multicellular layer, observed at
8:35 to 9:00. As cleavage proceeds, cell size indeed declines while the cell number
enhances leading to an advanced blastula in the limited blastodisc area. The blastoderm
happens to be opaque and firm. It also shows a broad and thick belt of the external yolk
syncytial layer. The animal surface of the yolk cell underlying the blastoderm is flat and
most of the yolk vacuoles are located inside. Shortly after, the periphery of the
blastoderm extends over the adjacent yolk margin (Fig. 36i).
3.3.2.4.1.2 The embryonic phase
Stage 10: epiboly, evacuation zone
Animal pole and vegetal pole are clearly separated from each other, this occurs around
16:00 h. The germ retakes on the circular shape with a lens-shaped blastoderm. The
upper part is densely packed with yolk. The blastoderm covers 30 % of the yolk surface
concurrently as the margin of the blastoderm is lying just beyond the lower margin of
the external yolk synctial layer. The inflated evacuation zone is covered by a single thin
and clear mono-layered epidermal enveloping layer. The animal surface of the yolk
sphere then turns out to be flat. The internal yolk syncytial layer gets more significant
prominent beneath the extremely slim and translucent blastoderm (Fig. 37a, 19:11).
Stage 11: 50% epiboly
The blastoderm prolongs spreading and covering 50 % of the yolk surface. As the
blastoderm extends vegetally, the external yolk syncytial nuclei moves underneath the
blastoderm to distribute over the yolk mass. The margin of the blastoderm can be
distinguished from the germ ring (Fig. 37b, 25:15).
Stage 12: embryonic shield
The margin of the blastoderm spreads over the yolk margin and a visible embryonic
shield emerges as a slender bulge. This longitudinal axis of the embryonic shield will be
later on recognized as the prospective embryo. The evacuation zone seems to be
dislocated from the vertical axis of the yolk mass (Fig. 37c, 27:00).
Stage 13: early neurula
The embryonic shield happens to be elongated towards the animal pole. It is formed by
a thickening of the ectodermal epithelium to form a classical neural plate. A small
evacuation zone is still present, but is not seen again after this stage (Fig. 37d, 29:15).
43
Fig. 36. Cleavage in Notopterus notopterus and blastulation. [a] Stage 1: one cell (zygote), 1: 10.[b]
Stage 2: two cells blastomeres (b), at 2:00 [c] Stage 3: four cells, 2:20. [d] Stage 4: eight cells, 2:42. [e]
Stage 5: early morula with 16 cells, 3:45. [f] Stage 6: asynchronous blastomeres up to circa 32 cells, 4:30.
[g] Stage 7: blastula with compact blastodisc (bd), 6:25 [h]. Stage 8: flat blastula, note the roundish
mound of the blastoderm on the top of the yolk, 7:40. [i] Stage 9: late blastula, the surface of blastoderm
appears smooth but the cells are still distinct, 8:35. Scale bar = 1 mm.
Stage 14: 75% epiboly
The blastoderm has almost covered up to 75 % of the yolk surface by the end of this
stage. The neural folds of the prospective head region are elevated from the epidermal
yolk sac cover. A notochord can be seen at the midline of the neural plate. This stage
morphologically classifies as rostral and caudal axis of the future embryo. The first
three somites are also visibly recognizable from this stage onward (Fig. 37e-f, 36:00).
Stage 15: wedge-shaped neural plate
The blastoderm expands spreading and covering up to almost 90 % of the yolk surface,
with yolk plug obviously much reduced from the previous stage. The neural plate
continues to extend laterally and becomes wedge-shaped (Fig. 37g, 39:10).
44
Fig. 37. Continuation of epiboly and neurulation in Notopterus notopterus. [a] Stage 10: evacuation zone
(ez) at animal pole and the deepening of the marginal zone (arrow) between the animal pole and the
vegetal pole, 19:11. [b] Stage 11: 50 % epiboly, germ ring (gr) positioned in between of animal pole and
vegetal pole, 25:15. [c] Stage 12: embryonic shields on the animal pole, note the translucent evacuation
zone (ez), 27:00. [d] Stage 13: onset of neurulation with vertical view, neural plate (npl), 29:15. [e] Stage
14: 75 % epiboly on vertical view, note the yolk plug is mostly constricted by the flat and short germ ring
(gr), 36:00. [f] Stage 14: transversal view, note the rostral (r) and caudal (c) part can be distinguished. [g]
Stage 15: 90 % epiboly; note the reduced yolk plug (yp), 39:10. [h] Stage 16: latest epiboly, note the tiny
remnant of yolk plug (yp), vertical view. [i] Stage 16: transversal view showing the wedge-shaped neural
plate (arrowhead), 41:35. Scale bar = 1 mm.
Stage 16: latest epiboly
The blastoderm covers almost the entire yolk surface, leaving an exposed yolk plug.
The germ ring is now swollen all around. The primary yolk sac cavity extends further
rostrally and caudally underneath the entire neural plate to form the segmentation cavity
(Fig. 37h-i, 41:35).
45
Fig. 38. Embryonic development in Notopterus notopterus. [a] Stage 17; spoon shaped, 48:50. Note the
the head cavity. [b] Stage 18: early trunk-tail bud, note the remnant of evacuation zone, 51:05. Note
somites (som) and the tail bud (tb) on the posterior part. [c] Stage 19: tail-bud bent, Kupffer’s vesicle
(Kv) appears in the bent tail bud, 84:00. Note the corpus cerebelli (cc) around the rhombencephalon area.
[d] Stage 20: otic placode (op) and heart beats (h) observed, 92:25. [e] Stage 21: fin-fold stage, 95:25.
Note the lobi linea literalis (lll). [f] Stage 22: eye pigment (e), 122:05. Note the broader fin fold (ff) along
the body. [g] Stage 23: “C” shaped, pre-hatching embryo, note the edge of the caudal fin, 146:50. Scale
bar = 1 mm.
46
Stage 17: spoon-shaped
Epiboly is completed and the yolk mass is wholly wrapped with both the blastoderm
and yolk syncytial layer. As neurulation proceeds, a neural groove becomes evident
along the midline of the plate, with a widened, spoon-shaped depression at the anterior
end. Shortly after the neural folds have approached each other along the midline, which
causes the neural plate to take a dumbbell-shaped appearance. In the prospective head
region, the neural folds thicken (Fig. 38a, 48:50).
Stage 18: early trunk-tail bud
The lateral edges of the neural plate are prominent and fold in toward the midline of the
embryo, mainly in the prospective trunk-tail region. The majority of caudal portion of
the neural anlage and underlying mesoderm project from the epidermal yolk sac cover
form a rudimentary trunk-tail region. The embryo now has around 10–13 somites
already. The earliest somatic muscular activity occurs in the posterior trunk-tail region.
The observed contractions were mostly weak lateral trunk-tail flexions. A first twitch
from touching the embryo was observed, which was mostly followed by a secondary
reaction or several trashes of the tail (Fig. 38b, 51:05).
Stage 19: tail bud-bent
This stage is also categorized by a 90° bend of the free trunk-tail. The somites and the
mesoderm destined for the further differentiation of somites is fused at the midline
above the spinal cord, and the mesodermal matter of the lateral plates in the head region
grows dorsally. The embryo now has 19–20 somites. The observed contractions were
mostly weak lateral trunk-tail flexions. Any actions from nearby will apparently
provoke a particular twitch, which is then followed later on by a secondary backlash or
even several thrashes of the tail. The straightening of the embryo enables then to
analyse the blood circulation in more detail. The straightening of the embryo now
allows following the blood circulation in more detail. The heart tube is currently visible.
The caudal tail region of the embryo, in which Kupffer’s vesicle is located, is separated
from the yolk sac and curved downward along it (Fig. 38c, 84:25)
Stage 20: otic placode and heart beat
The otic placode emerges for the first time at this stage. The yolk sac is covered by a
dense plexus of anastomosing subintestinal venae vitellinae. Spontaneous, vigorous,
flexing movements of the body are common, which cause the embryo to rotate within
the egg envelope. If illumination is changed, contraction frequency is shortly increased.
47
The central nervous system lays in a deep groove formed by the remarkably elevated
paraxial mesoderm in the trunk-tail region and the prominent, densely packed
mesoderm mass in the head region. The forming heart is seen inside the extended
pericardial cavity. The blood cells were circulating through a system which included in
a sequential order: a two chambered heart, a dorsal aorta from the heart to the posterior
limit of the yolk, a short intestinal loop, a subintestinal-vitelline vein to the oil globule
periphery, and after taking a diffuse random course across the external surface of the oil
globule back to the heart. The lobi lineae lateralis becomes visible (lll)
(Fig. 38d, 92:25).
Stage 21: fin-fold
The first sign of the embryonic fin fold is observed here. It runs along the midline from
the first somite caudally and encircles to the entire trunk-tail end. The trunk and tail
region constantly perform thrashing movements. There are 28 somites visible at this
stage. A dorsal aorta is discernible in the anterior body region. The entire head region
has significantly enlarged and shows several new features. A lens is formed inside the
eye cup, when the choroid fissure is still wide open. No pigment is discerned in the eye.
The yolk sac is covered by a dense plexus of anastomosing subintestinal venae
vitellinae. The membranous embryonic fin fold becomes conspicuous during the
illumination (Fig. 38e, 95:25).
Stage 22: eye pigment
Diffuse pigmenting appears in the eye. A few small unbranched melanophores that
contain diffuse melanine pigments are distributed all over the ectoderm of the head. The
muscular contractions of the embryo have proceeded to powerful movements of the
entire body by now. The vascular fin fold network extends over the entire fin fold,
covering the yolk sac. Some larger venae vitellinae are established at the lateral region
of the yolk mass surface. 48-52 pairs of somites could be observed (Fig. 38f, 122:05)
Stage 23: pre-hatching
The yolk cavity is enlarged beyond the entire anterior part of the embryo. The anterior
part of the head is slightly undercut at its juncture with the epidermal yolk sac cover. At
quiescent conditions the tip of the curved trunk-tail region reaches the vertical end of
the head. This tip may touch or even pass over the head during movements. The
embryos kept performing strong vigorous rotations and quivering motions. These
movements are sufficient to change the orientation of the embryo inside of the egg
envelope. The two columns of the presumptive corpus cerebelli fuse dorsally in the
midline, forming also a transversally widened bulge behind the fissure rhombo-
48
mesencephalica. This bulge curves forwards, inwards, ventrally and also grows
caudally, forming the primitive cerebellar cavity. The lobi lineae lateralis are very
obvious and start to elevate. The embryos are in a twisted position due to their
elongated body (Fig. 38g, 146:50).
3.3.2.4.1.3 The eleutheroembryonic phase
Stage 24: hatching
Hatching started at about 168-204 hrs after spawning and the free embryos measured
about 10.5 mm. The egg envelope had lost its stability before, which could be seen by
its pliable texture. The tail usually protrudes first and usually after a part of the egg
envelope ruptured; the free embryos with the head part often still remain within the egg
envelope for some time. The yolk sac measured around 3.8 mm in total length at this
stage, situated mostly still inside the egg envelope. As the upward fin fold presents a
membranous bud of the dorsal fin, the development of the dorsal fin can be followed
from this stage onward (Fig. 39a, 168:05).
Stage 25: completed rupture
The egg envelope has completely ruptured. It took around 20 minutes until the embryo
successfully detached from its egg envelope. Shortly after hatching, the embryos rest
calmly and in a straight line on the ground. The eyes are completely black. The tiny,
hemispherical pectoral fin projects from the dorsal epidermal yolk sac cover, and is
positioned close to the heart region. The yolk sac still measures around 3 mm. There are
58–62 pairs of somites observed at this stage (Fig. 39b, 168:25).
Stage 26: jaw and branchial placode
The head still remains bent downwards along the yolk sphere. The first mesenchymal
condensations of jaws and branchial arches can be seen between head and yolk mass.
Usually the yolk mass is no longer rounded, but has become egg shaped with a narrow
posterior end. The yolk sac measures around 2 mm and the free embryos usually
measure 11 mm at this stage. When the freshly hatched embryo is stimulated, it makes
rapid, striking movements, especially with its trunk and tail, through which it whirls
around in circles. The early free embryos are still too heavy for directed movements
(Fig. 39c, day 8).
49
Fig. 39. Hatching process and free embryonic in Notopterus notopterus. [a] Stage 24: just hatched-after
breaking the weak egg envelope, caudal part has relieved, anterior part still attached to the remnant of the
egg-envelope, note the emergence of dorsal and anal fins. Yolk sac (yc) still covered with egg envelope,
168:05. [b] Stage 25: rupture the complete egg-envelope and free anterior part of the embryo, 168:25.
Note the tiny pectoral fin bud, black arrowhead. [c] Stage 26: jaw and branchial placode, day 8. [d] Stage
27: mouth opening, day 10. Note the emergence of swim bladder (sb), arrow. White arrowheads depict
gradual development of dorsal fin. Scale bar = 5 mm
50
Stage 27: mouth opening
Some evident changes in the outer appearance and in the structure of the head can be
easily seen. The head process has already undergone a significant lift. Several structures
are clearer defined. In particular, the heart is beginning to descend into the pericardial
cavity. The anlagen of the dorsal and anal fins appear simultaneously as denser
concentrations of mesenchyme in the fin fold. The formation of the first rays in the
caudal fin is evident from this stage onward. The future swim bladder can be pointed
out. Melanophores spread out significantly not only on the head part but also appear for
the first time on the trunk and tail part. The head lift and further growth and
straightening of the head exposes the underside of the head.
The lower jaw has straightened forward. Mouth opening shows a “>” shaped with clear
separation between upper and lower jaws. The central lepidotrichial rays began to
segment. The yolk sac measures around 1.4 mm and the embryos measure 12 mm
(Fig. 39d, day 10). The tiny pectoral fin bud becomes ellipsoid in shape due to its
dorsocaudal prolongation, as seen detail in Figure 40. The pectoral fin is clearly larger
than the eye now.
Fig. 40. Pectoral fin buds (pfb) in Notopterus notopterus, larger magnification - day 9. mmesencephalon; lj- lower jaw; nch – notochord; Ao – aorta dorsalis; Vsi – vena subintestinalis; Vc – vena
caudalis; af – anal fin; g- gils; h – heart; lev – left efferent vena vitellina; ys – yolk sac. Scale bar = 2 mm.
51
Stage 28: commence on median fin-fold regression
The dorsal and anal fins lengthen over the embryonic fin fold. The continued regression
of the median fin fold allows determining the contour of the dorsal, anal, and caudal
fins. The pectoral fin anlage now is circular with an enlarged and indented rim. The
haemal and hypural processes are covered by a few muscles. From this stage onward,
the embryos become increasingly more mobile. The embryo is now capable of
completely lifting its head and actively closing or opening its mouth. The upwards
flexion of the end of the notochord has reached its final shape, perpendicular to the
longitudinal axis of the embryo. The embryos measured 13.6 mm and the capacity of
the yolk-sac had decreased significantly (Fig. 41a, day 12).
Stage 29: late embryonic
The embryonic fin fold has significantly shrunk in height, especially in the dorsal part.
The embryos measured 14.9 mm. The yolk sac remains visible. The pectoral fins are
now fully formed with 6 segmented lepidotrichial rays each. They are functional and
used as in other teleostean larvae for propulsion. In this stage, first movements of active
breathing of the gill cover were observed (Fig. 41b, day 14).
3.3.2.4.2 The larval period
Stage 30: exogenous feeding
The first stage of the larval period is characterized by exogenous feeding concurrent
with endogenous nutrient-utilization. The yolk sac remnant is still present and almost
absorbed.The swim bladder appears tube-shaped and continues to develop from this
stage onward. The dorsal fin has evolved showing newly developed fin-rays. The final
shape of the caudal fin is reached by the conspicuous constriction and complete
regression of the remaining fin fold at the caudal end of the peduncle. Melanophores
spread over the exposed integument. Pectoral fins are well developed and already
mobile. The depicted individual larva at this stage measures 16.2 mm (Fig. 41c,
day 17).
Stage 31: developed-eye
The swim bladder starts to elongate dorso-rostrally in order to connect with the otic
vesicle. The yolk sac remain now completely absorbed. From this stage onward, the
larvae are very mobile and actively hunting for higher food densities. The ring of the
eye has developed well and the larvae start to swim, although only on the bottom. The
swim bladder has elongated a bit more and appears thinner. An intensely yellow mass
52
filled the gut. The total length of the depicted individual larva is 21 mm (Fig. 41d, day
24).
Fig. 41. Late embryonic and larval development in Notopterus notopterus. [a] Stage 28: pronounced
regression of median fin fold, day 12. Note almost complete reduced yolk sac (ys). Note the pectoral fin,
arrow. [b] Stage 29: late embryonic phase, day 14. [c] Stage 30: developed-swim bladder, day 17. [d]
Stage 31: pigmented iris is very distinct, day 24. [e] Stage 32: caudal lobe separation, day 36. Arrowheads
depict gradual development of dorsal fin. Scale bar = 5 mm.
53
Stage 32: caudal lobe separation
The dorsal and anal fins separate from the peduncular fin fold remain and the
asymmetry of the caudal fin has become more conspicuous. The larvae usually measure
24 mm or more at this stage (Fig. 41e, day 36). The density of melanin has been
multiplied on the entire body, especially underneath the pectoral fins and the area above
the intestine. The development of the dorsal and caudal fin is presented largely
magnified and in detail in Figure 42 and 43 below.
Fig. 42. Sequence of the development of the dorsal fin starting from hatching until the end of larval
period in Notopterus notopterus. [a] shortly after hatching, day 7. [b] Dorsal fin bud slightly appeared,
day 8. [c] Distinct dorsal fin and regressing of fin fold, day 11. [d] Fin rays on dorsal fin can be seen, day
12. [e] Well-developed dorsal fin with 9 fin-rays, 17 days. [f] Dorsal fin in a juvenile, 36 days. White
arrowhead depicts the gradual development of dorsal fin. Scale bar = 2 mm.
3.3.2.4.3 The juvenile period
The juvenile period of Notopterus notopterus comprises five stages as follows:
Stage 33: juvenile – 1
This stage can be clearly characterized as a transition mark from the larval period to the
juvenile period by its profound change in the pigmentation pattern. The embryonic fin
folds are no longer visible.The swim bladder elongates further and is densely covered
with melanophores. The lateral line of the body becomes visible, starting right behind
the head region at this stage. The formation of tube-like anterior nostrils is also starting
from this stage onward. The body grows significantly in height and melanophores
become denser also in the caudal part. Silver-coloured scales appeared on the midline of
54
the anterior part of the trunk and at the base of the pectoral fin and caudal fin. The
expansion of the stomach and the intestine can be clearly seen. The typical early
juvenile measures now 29 mm (Fig. 46a, day 52).
Fig. 43. Sequence of caudal fin development starting from hatching until larval period in
Notopterus notopterus. [a] shortly after hatching, day 7. [b] Haemal and hypural plate (hh) emerged. Note
the slight regression of fin fold, day 10. [c] Completely disappeared fin fold with 10 caudal fin rays, day
20. [d] Well-developed caudal fin in a juvenile, day 36. cfm- caudal fin mesenchyme; ff- fin folds; cfr –
caudal fin rays; cf – caudal fin; af – anal fin. Scale bar = 2 mm.
Fig. 44. Two sequences of female’s genital papilla in Notopterus notopterus in comparable magnification.
[a] The emergence of genital papilla in female with total length 52 mm. [b] Genital papilla of female with
total length of 145 mm. Note the significant growth of genital papilla (white arrowhead) in between
ventral fins (arrow) and anal fin. Scale bar = 2 mm.
55
Fig. 45. Complete development of female’s genital papilla in Notopterus notopterus, seen in various
magnified photographs. [a] The emergence of genital papilla, total length (TL) 52 mm and total weight
(TW) 1.77 g, day 83. Note the developed pair of ventral fins. [b] Female’s genital papilla with TL 100
mm, 5 months. [c] TL=124 mm, 8 months. [d] TL= 136 mm, 12 months. [e] TL=145 mm and TW= 28.5
g, in 18 months old. [f] Mature genital papilla obviously longer than the pelvic fins, TL= 230 mm,
captured at 24 months old. Note the significant growth of genital papilla (white arrowhead) in between of
ventral fins (arrow) and anal fin. Scale bar= 2 mm (a, c, e); 5 mm (b, d, f).
Stage 34: juvenile - 2
At this stage the body has fully changed its colour into vertically arranged dark brown
stripes. There is a slight black colour at the outer part of the eye. This typical juvenile-2
measures 34 mm (Fig. 46b, day 70)
Stage 35: juvenile - 3
The body shows much stronger dark brownish stripes and commencing elongation of
tubular nostrils is apparent at the snout. The genital papilla emerges for the first time.
The depicted individual measures 46 mm TL (Fig. 46c, day 80).
56
Stage 36: juvenile - 4
There is once again an obvious change in the pigmentation pattern from the previous
stage. The striped, dark-brown colouring has disappeared. The whole body turns into
brownish-gold and scales are externally visible from this stage onward. The pale
remains of dark brown stripes can still be recognized. The pelvic fins start to emerge at
this stage and the depicted individual measures 76 mm TL (Fig. 47a, day 92). Series
development of genital papilla in a female is shown in Figure 44 and 45.
Stage 37: juvenile - 5
The dorsal fin looses its trapezoidal shape and forms into the triangular shape of the
adult. The specimens have attained the general body profile of the immature adult now.
The genital papilla is clearly recognisable behind the elongated pelvic fins. The depicted
late juvenile individual measures 145 mm TL (Fig. 47b, 18 months).
3.3.2.4.4 The adult period
Stage 38: adult
The head shape and body pigmentation are different from the late juvenile. From this
stage onward, the genital papilla of the mature can be easily differentiated between male
and female by its form. The female’s genital papilla is whitish, broader and shorter,
whereas the male’s is yellowish, thinner and longer. Gonad maturation can be easily
recognized by the swollen abdomen and increase of appetite starting at the age of 24
months. The illustrated individual adult measures 275 mm TL (Fig. 47c, 30 months). A
glance of all development stages in Notopterus notopterus is presented in Table 6.
57
Fig. 46. Juvenile transformation in Notopterus notopterus. [a] Stage 33: early juvenile in larger
magnification, note the appearance of lateral line and short tubular nostrils, day 52 [b] Stage 34: stripe
colored body of juvenile, day 70. [c] Stage 35: strong stripe coloured alongside the body, note developed
tentacles of the anterior nostrils, day 80. Arrow points the base of well developed pectoral fin.White
arrowheads show the anterior nostrils. Black arrowhead points the genital papilla. Scale bar = 10 mm.
58
Fig. 47. Late juvenile to the maturation stage in Notopterus notopterus. [a] Stage 36: appearance of
ventral fin, black arrowhead, day 92. [b] Stage 37: late stage of juvenile with total length of 145 mm, note
the change in colour and the emergence of scales, 18 months. [c] Stage 38: adult male with total length
275 mm, captured at age of 30 months. Note the ventral fin (arrowhead) longer than the genital papilla
Scale bar = 10 mm (a); 20 mm (b); 30 mm (c).
59
Table 6. Overview of developmental stages of Notopterus notopterus (27 °C). Determination of periods
after Balon (1975)
Period:
Embryonic
Phase
Stage
Age
(hrs:min)
Characteristics
Figure
Cleavage
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
1:10
2:00
2:20
2:42
3:45
4:30
6:25
7:40
8:35
19:11
25:15
27:00
29:15
36:00
39:10
41:35
48:50
51:05
84:00
92:25
95:25
122:05
146:50
168:05
168:25
day 8
day 10
day 12
day 14
day 17
day 24
day 36
day 52
day 70
day 80
day 92
18 months
30 months
Zygote-1 cell
2 Blastomeres
4 Blastomeres
8 Blastomeres
Early morula
Late morula
Blastula
Flat blastula
Late blastula
Evacuation zone
Epiboly 50%
Embryonic shields
Early neurula
Epiboly 75%
Wedge-shaped neural plate
Latest epiboly
Spoon-shaped
Early trunk-tail bud
Tail bud bent
Otic placode and heart beat
Fin fold
Eye pigment
Pre-hatching stage
Hatching stage
Completed rupture
Jaw and branchial placode
Mouth opening
Emergence of finfold regression
Late embryonic
Swim bladder
Developed eye
Caudal lobe separation
Juvenile 1
Juvenile 2
Juvenile 3
Juvenile 4
Juvenile 5
Adult
36a
36b
36c
36d
36e
36f
36g
36h
36i
37a
37b
37c
37d
37e-f
37g
37h-i
38a
38b
38c
38d
38e
38f
38g
39a
39b
39c
39d
41a
41b
41c
41d
41e
46a
46b
46c
47a
47b
47c
Embryonic
Larval
Eleutheroembryonic
Protopterygiolarval
Pterygiolarval
Juvenile
Adult
60
3.4 Osteoglossidae
3.4.1 Osteoglossum bicirrhosum
3.4.1.1 External morphology and in situ condition of dissected gonads
Figure 48 and 49 illustrate the Silver Arowana (Osteoglossum bicirrhosum) and Blue
Arowana Osteoglossum ferreirai, respectively. Male and female of both species do not
show a clearly visible sexual dimorphism. The female can only be recognized by its
swollen belly during the maturation phase. Osteoglossum ssp. has a pair of barbels
located on the lower jaw. In both species strong and hard cycloid scales cover the entire
body.
Fig. 48. A female of Osteoglossum bicirrhosum [a] of 63 cm total length and 59 cm standard length.
[b] In the head the diameter of the eye is indicated as 18 mm, the characteristic barbells, and a large scale
taken out from the dorsal part of the body. [c] The posterior part with caudal, anal and dorsal fins in gold
or brown. [d] In situ condition of dissected abdomen. [e] Single ovary on the left side of the body cavity
is densely covered by fat, situated on the right corner of the body cavity, arrow. Scale bar = 10 cm.
61
The silvery-brown Osteoglossum bicirrhosum (Fig. 48a) shows an elongate body
covered with large scales (Fig. 48b) as well as the long anal fin together with the caudal
fin and dorsal fin (Fig. 48c). The barbels comprise 20 mm in length in the shown adult
specimen (Fig. 48d). Osteoglossum ferreirai (Fig. 49a) is similar in external
morphology to O. bicirrhosum, except for the anal fin, caudal and dorsal fins, which are
blue (Fig. 49b, c). The barbels of the Blue Arowana, presented in Fig. 49d, are 18 mm
in length. In situ conditions (Fig. 48e, 49e) of female specimens of both species reveal
one single ovary, which is located in the left hind corner of the body cavity. Among 16
specimens (11♀, 5♂), total gonad weight (TW) of the females ranged from 1.7 to 48.1
g, with total length (TL) varying from 51.5 to 67 cm, whereas total weight of the males
testes varied from 0.3 to 0.7 g, with TL ranging from 40.2 to 60 cm.
Fig. 49. A female of Osteoglossum ferreirai (Blue Arowana). [a] 60 cm total length and 56.5 cm standard
length. [b] Blue colour of anal, [c] caudal and dorsal fin can be clearly seen. [d] In situ condition of
opened abdomen. [e] Ovary is filled mostly with stadium IV oocytes. Scale bar = 10 cm.
62
Fig. 50. Two ovaries of Osteoglossum bicirrhosum. [a] with total gonad weight (TW) 31.5 g and [b]
Osteoglossum ferreirai with total gonad weight 39.8 g. Both ovaries clearly contain large numbers of
oocytes in different stages of development accompanied by some fat, arrow. Photos were taken shortly
after dissection and measurement of gonad weight. Scale bar = 2 cm.
3.4.1.2 Gonad of Silver Arowana and Blue Arowana
The gonads of dissected O. bicirrhosum (Silver Arowana) and O. ferreirai (Blue
Arowana) are illustrated in Figure 50. The ovaries were filled with oocytes in different
stages (stadium I to stadium IV). Maturity coefficients (MC) of these two ovaries are
2.75 % (O. bicirrhosum, TL = 63 cm) and 3.4 % (O. ferreirai, TL= 60 cm),
respectively.
Fig. 51. [a-d] Freshly collected non-adhesive eggs of Osteoglossum bicirrhosum. Two different colours
of eggs, [a, b] yellow and [c, d] orange, were found during this study of eggs with the same diameter of
12 mm. The eggs with transparent chorion, [a, c] viewed from the side of a Petri dish and [b, d] others
viewed from the top. Scale bar = 2 mm.
63
3.4.1.3 Egg of Silver Arowana
Table 7 presents the list of visited farms of Silver Arowana within five months
including the numbers of collected, fresh eggs, numbers of collected eggs with embryo
and numbers of collected juveniles with the yolk sac, taken directly from the male’s
mouth. Egg numbers per collecting varied from 23–220. The eggs of O. bicirrhosum
ranged from 11 to 12 mm in diameter with a total weight of 0.9–1.08 g. They are
roundish, non-adhesive and easily sink to the bottom (Fig. 51). The eggs are covered
with a thin, transparent and fairly robust egg envelope, which encloses a narrow
perivitelline space. As figure 51a and 51c illustrate, there were two different colours of
eggs found: orange and yellow. The phenomenon of yellow coloured eggs was mostly
found in female specimens younger than 28 months, whereas in specimens older than
28 months the gonads contained mostly orange coloured eggs.
Table 7. All visited farms of Osteoglossum bicirrhosum in Florencia and other cities nearby together with
the numbers of collected fresh eggs (♂1), eggs containing embryo (♂2) and juveniles with yolk sac
(♂3 and ♂4) taken directly from male’s mouth
No
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
Name of Farm
Hermes
Navaja
Diego Palace
Luis Helga
Christian
Ricarvte Silva
David Garcia
Navaja
Navaja
Navaja (09:30 am)
Navaja (17:25 pm)
Navaja
Luis Helga
Diego Palace
Alex
Navaha
Viktor
Doncello-Acuica
Hermes
Hermes
Navaja
Diego Palace
Victor
Date
14.01.2011
16.01.2011
19.01.2011
21.01.2011
27.01.2011
30.01.2011
01.02.2011
02.02.2011
09.02.2011
19.02.2011
19.02.2011
22.02.2011
28.02.2011
12.03.2011
13.03.2011
22.03.2011
28.03.2011
17.04.2011
24.04.2011
24.04.2011
01.05.2011
14.05.2011
16.05.2011
♂1
0
0
0
45
0
0
0
220
0
27
23
0
0
0
40
145
96
0
112
106
0
0
25
♂2
0
0
0
10
0
0
0
72
0
103
21
0
0
0
6
0
0
31
0
0
0
0
0
♂3
0
0
0
0
0
0
0
0
0
80
0
0
0
0
0
0
0
35
0
0
0
0
0
♂4
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
33
0
0
0
0
0
64
3.4.1.4 Development
The exact time of fertilization is unknown since the eggs were collected directly from
the male’s mouth. Therefore the age given in this study was calculated from the time,
when the eggs had been collected in the farm. Eggs and embryos were incubated and
raised at 28 °C. The terminology of the development of Osteoglossum bicirrhosum
follows the classifications of Balon (1975). There are five periods: the embryonic period
(I), the larval period (II), the juvenile period (III), the adult period (IV) and the
senescent period (V The larval period will not be presented, since most likely this
species does not undergo this particular period. Similar to Notopterus notopterus the
adult period is also excluded in this work. Time of egg collection is defined as time zero
in age determination; it is presented as hrs:min or days.
3.4.1.4.1 The embryonic period
3.4.1.4.1.1 The embryonic phase
Stage 1: freshly captured eggs
Newly spawned eggs were observed circa 3:00 after collecting the eggs from the farm.
The eggs are non-sticky, translucent, and roundish in shape and possess a large yolksac. Due to its large yolk it is really difficult to position the egg properly (Fig. 52a).
Stage 2: neural plate
Much later cleavage and gastrulation is followed by neural plate formation at 8:00,
which extends laterally and becomes (Fig. 52b) a distinct neural plate observed as seen
in Figure 52c. A trunk tail mound was evident almost at the same age. There was an
aggregation of cells in the head area.
Stage 3: onset somatogenesis
As neurulation continues, a neural groove becomes obvious and lengthens along the
midline of the plate, with a widened, spoon-shaped head formation at the anterior end.
The location of the future eye can be seen. The head starts to develop at this stage. Brain
regionalization, a lateral symmetrical thickening of the neural folds, appears in the head
region. The head area was swollen, showing first indications for developing brain parts.
It filled the perivitelline space and touched the membrane next to the micropyle. The
presumptive heart tube is already positioned on the right base of the anterior of the
body. No blood elements were visible. The neural plate is longer and broader than in
previous stage (Fig. 53a-c, 21h).
65
Fig. 52. First stages of the embryonic phase in Osteoglossum bicirrhosum. [a] Stage 1: freshly captured
eggs, around 3 hours after released from male’s mouth. [b] Stage 2: embryonic development with the
axial trunk-tail (arrow) and anterior part of the body, future head located (arrowhead), 8 hours.
[c] Depicting figure b with magnified 5x larger showing clearly embryonic development comprising
anterior part (arrowhead), neural plate and the trunk-tail mound (t). Scale bar = 3 mm.
Stage 4: brain regionalisation
There were plenty of red speckled pigments on the yolk as a first sign of blood
circulation. Head region enlarged due to further development of the brain region. The
anterior part shows the future upper and lower jaw. The heart became visible and beat
regularly. The evacuation zone in comparison to the previous stage has become much
larger. This occurred 36 hours after releasing the egg from male’s mouth (Fig. 54a-b).
66
Fig. 53. Continuation of embryonic development in Osteoglossum bicirrhosum, stage 3 (a-c). [a] Around
21 hours after being released from male’s mouth, view from front side. [b] View from front part of head
region; note the beginning of heart and a pair of lens (l). [c] View from dorsal showing the neural plate
(arrow). Scale bar = 3 mm.
Stage 5: early trunk-tail bud
At around 42 hours, a mass red blood cells and the forming heart can be seen inside the
extended evacuation zone. The neural plate stretches rostrally and caudally now beyond
the epidermal yolk-sac cover. Most of the caudal region of the neural anlage is now
forming a rudimentary trunk-tail bud. Figures 55 (a-d) illustrate four different
positions; embryo with 7 mm in total length still inside the egg envelope, thrashing
67
movements occurred as normal activity of the trunk and tail region. The eye lens and
enlargement of the head can be seen here. Blood flow through the hepatic anlage that
produced the hepatic-vitelline vein on the left anterior dorso-lateral surface of the yolk.
Fig. 54. Continuation of embryonic development in Osteoglossum bicirrhosum, stage 4. [a] The embryo
36 hours after release, showing massive red spots known capillaries on the entire yolk as first sign of
blood circulation. [b] View from front part, showing enlarged head region. Note the heart beat, white
arrowhead. Scale bar = 3 mm (a), 1 mm (b).
Stage 6: neural anlage
After 56 hours, the neural folds become contiguous along the dorsal midline of the
trunk-tail, later on become fused to form a neural rod. Increased numbers of capillaries
are seen on the entire yolk-sac and an opaque vena vittelina in direction to heart can be
seen. If illumination is changed, contraction frequency is temporarily increased
(Fig. 56a-d).
Stage 7: pectoral fin bud
The earliest somatic muscular activity occurs in the posterior trunk-tail region.
Touching the embryo will first elicit a single twitch, then later a secondary backlash or
even several thrashes of the tail. The neural plate projects rostrally and caudally now
beyond the epidermal yolk-sac cover and is now formed like a club. A lateral
symmetrical thickening of the neural folds suddenly emerged in the head region. At this
stage the blood elements start circulating. The embryos are now 78 hours old and
measure 10–12 mm in total length. Pectoral fins appear for the first time at this stage.
Four branchial arches can be recognized behind the eye and above the heart. Only a thin
layer of yolk separates the stomach from the ventral epidermal yolk sac cover
(Fig. 57 a-b).
68
Fig. 55. Continuation of embryonic development in Osteoglossum bicirrhosum, Stage 5. [a] The embryo
42 hours after release, reduced number of red spots and first recognition of vena vitellina in direction to
heart. [b] Left lateral view, showing the completely lengthened body with a lot of mesenchyme around
the trunk tail. [c] Head region and the whole body [d] with trunk tail bud (tt) in larger magnification.
Arrowhead shows the position of heart as orientation. Scale bar = 3 mm (a, b, d); 1 mm (c).
Stage 8: pre-hatching
The pre-hatching embryo appears with large black eye and gills at 160:00 (Fig. 58a).
The pectoral fin bud becomes ellipsoid in shape, by dorsocaudal prolongation, and is
now as big as the eye. The first formation of the rays in the caudal fin is now visible.
The muscular contractions of the embryo are such powerful actions that they affect the
position of the entire body. These strong movements are indeed sufficient to change the
orientation of the embryo inside the egg envelope. Later on, the embryo will start to
break the egg envelope gradually with its tail which causes the egg envelope to lose its
stability (Fig. 58b).
69
Fig. 56. Continuation of embryonic development in Osteoglossum bicirrhosum. [a-d] Stage 6: embryo in
56 hours after release in various positions, [a] front part, note the heart and the red vitelline vein (vv) in
direction to the heart, [b] back part, note the trunk tail (tt) mound, [c] transversal, midline region of the
neural plate with formed notochord (nch), and [d] view from dorsal, shows the entire body. Note active
heart beat (h). Scale bar = 3 mm.
Stage 9: hatching
Hatching occurred at 162-166 h (around 7 days) after being released from parent’s
mouth. Shortly after rupture of the egg envelope, the free embryos often stay inside the
egg envelope for a while (Fig. 58c). It took around 20 minutes releasing its complete
body from the egg envelope. The mouth was not yet open but was completely formed
and the heart is located on the ventral side of the branchial area. The yolk was covered
with denser capillaries than previously. Now the embryos are undergoing a more
complex blood circulation process and they measure around 16 mm (Fig. 58d).
70
Fig. 57. Continuation of embryonic development of Osteoglossum bicirrhosum, Stage 7. [a] view from
front part and [b] elongation of the trunk-tail and the emergence of pectoral fin bud (arrow), 72 hours.
Scale bar = 3 mm
3.4.1.4.1.2 The eleutheroembryonic phase
The eleutheroembryonic will be characterized from stage 9 to stage 13, starting right
after hatching until the embryos start to feed externally with the yolk sac still attached.
Stage 10: free embryo
The free embryos measure 17 mm on day 8 and stay instable with body on the upper
part of yolk sac, though often also its bodies lie on the bottom of the tank. When
disturbed, the free embryos wiggled their trunk and tail, but still cannot move the whole
body with their large yolk sac. After release, embryo’s motion was erratic though still
moving forward. The pectoral fins were already capable of continuous movement and it
was as big as the eye. The branchial arteries already formed five pairs of arches. An
extensive band of melanophores appeared, covering the brain region. Embryos were
either lying down on their dorsal side in a stable state or hanging with their large yolk
sac unstably for a short time. The head structure is better defined (Fig. 59a, day 8).
Stage 11: jaw and branchial placodes
One to two days after hatching, the development of the circulatory system was
obviously more extensive. The mouth was not completely developed and still couldnot
be closed yet. Between head and yolk mass the first mesenchymal condensations of
jaws and branchial arches could be recognized. The head process had already undergone
a significant lift with several structures are being more defined. The branchial
circulation was strongly developed and the gill filaments began to vascularise. The
pharynx can be clearly seen through the transparent embryo. Its rostral edge attached to
71
the body, while the caudal edge sticks out from the body. The diameter of the yolk sac
measured around 12-13 mm. The embryo now has 36 somites and measure 18 mm
(Fig 59b, day 9). The alteration of head development from shortly after hatching at day
7 until day 9 is presented in Figure 60 in larger magnification.
Fig. 58. Pre-hatching and free embryonic development of Osteoglossum bicirrhosum. [a] Stage 8: embryo
with developed pectoral fin (black arrowhead). Note the strong red vena vitellina (vv). [b] Embryo started
to break through the egg envelope by assertively moving its tail, 160 h. [c] Stage 9: Hatching, the free
embryo is still attached to the remains of the egg envelope, view from left lateral (L). Note the pectoral
fin bud (white arrowhead), 162-166 h. [d] View from behind (B), note the branchial arches (black
arrowhead) and direction of blood circulation (black arrow). h – heart, vi – vena caudalis inferior. Scale
bar = 4 mm.
Stage 12: initiation of barbels
The pelvic fin bud is already formed as a membranous bud, at the anterior to the anal fin
and develops from this stage onward. Melanophores blacken at the edge of the lower
72
mouth; it’s the sign of the barbels formation. The lower jaw has elongated and
straightened. The mouth was entirely developed and moved rhythmically. The anlagen
of the dorsal and anal fins appear simultaneously as denser concentrations of
mesenchyme in the fin fold. The formation of the first rays in the caudal fin is distinct
now. From this stage onward, the mobility has increased. The depicted individual
measures 24 mm at this stage (Fig. 61a, day 12).
Fig. 59. Free embryonic development of Osteoglossum bicirrhosum. [a] Stage 10: Almost 15 hours after
hatching with completed rupture of the egg envelope. Note the direction of blood circulation to the heart,
day 8. [b] Stage 11: free embryo, 9 days. Note the ellipsoid shaped of pectoral fin (pf) with an open
mouth. h – heart, er – eye ring, t – telenchepalon, m – mesencephalon, Vsi – vena subintestinalis,
nch-notochord, Ao – aorta dorsalis. Scale bar = 3 mm.
73
Fig. 60. Alteration of head structure of free embryo from stage 9 to stage 11 in
Osteoglossum bicirrhosum, larger magnification. [a] Stage 9: the head is still bent downwards along the
yolk sac and black pigmentation around the forebrain can be seen, day 7. [b] Stage 10: head elongation
particularly on the posterior part, day 8. [c] Stage 11: head lifted with an open mouth, 9 days. Note the
ellipsoid shaped of pectoral fin (pf). Scale bar = 2 mm.
74
Stage 13: ventral fin bud and swim bladder
At this stage the embryos are very actively swimming although only for a short
distance. The swim bladder starts to elongate dorso-rostrally. At this stage, the pectoral
fins are completely formed and functional with their 6 segmented lepidotrichial rays.
Dorsal view of head region during this stage shows pectoral fins as two-equal wings.
The head and around the dorsal part are densely covered with black pigments. The oval
shaped yolk sac has gradually changed to spherical shape onward. The yolk was
covered with more capillaries than previously. The surface of the still large yellow yolk
sac was permeated by numerous finer capillaries than in the previous stage. Now the
eleutheroembryos reached 28 mm in total length and the diameter of yolk sac measure
12 mm now (Fig. 61b, day 15). A large number of embryos in this stage are presented
in Figure 62 showing embryos lying unstably on the ground.
Fig. 61. Starting the eleutheroembryonic phase of Osteoglossum bicirrhosum. [a] Stage 12: massive head
pigmentation, day 12 [b] Stage 13: developed fins, note the migrating of yolk into the abdominal cavity,
day 15. Scale bar = 4 mm.
75
Fig. 62. A mass of free embryos of Osteoglossum bicirrhosum with age of 15 days, taken from the
aquarium. [a, b] Embryos lie unstably on the bottom of the tank with little movement. Note the
pigmentation around the head region and caudal part.
Stage 14: late eleutheroembryo
The barbels elongate in shape and measure 1 mm. Silver colour intensifies around the
eye to the operculum region. A pair of ventral fins on the base part of the belly can be
seen from this stage onward. It sticks to the yolk sac cavity. Two long, black-pigmented
rows elongated dorso-rostrally and reached the anal fin. Melanophores covered almost
all part of pectoral fins and left the outer part white, as well as around the area of
operculum and the snout part. The late eleutheroembryos now measure 34 mm and the
diameter of yolk sac still measure 12 mm (Fig. 63a, day 24).
3.4.1.4.2 The juvenile period
Stage 15: juvenile - 1 (Mixed feeding)
The pectoral fins develop with dense black pigments on the edge of the fins. Opaque
melanophores covered the whole body with its particular aggregations at the top of the
head and with very dark dark rows along the dorsal midline, along the intestine and the
bases of the dorsal and anal fins. At this stage the juveniles start to feed exogenously
concurrently with endogenous nutrient-utilization. The juveniles measure 38 mm
(Fig. 63b, day 26).
Stage 16: juvenile - 2
The yolk sac has very much declined and the pelvic fin has developed further. Dense
network melanophores covered the head region and also two dark rows along the dorsal
midline and the centre part of the body to the caudal fin emerged. The barbels measure
2 mm and the juveniles measure 40 mm at this stage (Fig. 63c, day 30).
76
Fig. 63. [a] Continuation of the eleutheroembryonic phase and [b-d] starting point of the juvenile stage in
Osteoglossum bicirrhosum. [a] Stage 14: late phase of the eleutheroembryo, day 24. [b] Stage 15: first
juvenile, day 26. [c] Stage 16: a-30-day juvenile. [d] Stage 17: scales initiation, day 36. Scale bar =
10 cm.
77
Stage 17: juvenile - 3
The caudal part of yolk sac has released from the body due to the elongation of ventral
fin. Usually the yolk mass is no longer rounded as previous stages, but has become a
semicircle-shaped. Black pigmentation appears stronger along the body and the scales
commence to develop from this stage onward. The mouth is able to close completely at
this point. The pelvic fins are well-visible. The juveniles measure 45 mm at this stage
(Fig. 63d, day 36).
Stage 18: juvenile - 4
The pelvic fins reached 8 mm in total length and the barbels are 3 mm long. The yolk
sac has elongated quite a lot and has become tube-shaped. The depicted individual
measures 48 mm (Fig. 64a, day 40).
Stage 19: juvenile - 5
The pelvic fins measure 9 mm and the barbels are still 3 mm long. The yolk sac is
shorter than in the previous stage. The specimens now measure 56 mm
(Fig. 64b, day 50).
Stage 20: juvenile - 6
Yolk sac capillaries somehow reduced although still clearly recognizable as to their
origin. The barbels measure 4 mm and the pelvic fins measure 10 mm. The specimens
measure 60 mm at this stage (Fig. 64c, day 64).
Stage 21: juvenile - 7
The barbels measure 5 mm and the pelvic fins reached 13 mm in length. The yolk
almost disappeared within the body cavity walls, although a tiny rest of it remained. The
juveniles measure 80 mm (Fig. 64d, day 75)
Stage 22: juvenile - 8
The barbels measure 6 mm now and the elongate pelvic fin has reached 15 mm in length
at this stage. The body of the juvenile has elongated further and developed large silver
scales. The linea literalis shows a long line starting from behind the head region and
somehow with stripe line showing a 135° shaped. The juveniles now measure 125 mm
(Fig. 64e, day 100). A glance of all development stages in Osteoglossum bicirrhosum is
presented in Table 8 as below.
78
Fig. 64. Continuation of the juvenile sequences in Osteoglossum bicirrhosum. [a] Stage 18: a long tubeshaped yolk sac, day 50. [b] Stage 19: much reduced yolk sac. [c] Stage 20: tiny remnant of yolk sac, day
64. [d] Stage 21: juvenile prior fully absorbed yolk sac, day 75. [e] Stage 22: completely disappeared of
yolk sac also called as late juvenile state, day 100. Scale bar = 10 mm.
79
Table 8. Overview of developmental stages in Osteoglossum bicirrhosum (28 °C), Determination after
Balon (1975)
Period
Embryonic
Phase
Stage
Embryonic
1
2
3
4
5
6
7
8
9
10
11
12
Age
(hrs:min)
03:00
08:00
21:00
36:00
42:00
56:00
78:00
160:00
162:00–166:00
D6–D7
D8
D12
13
14
15
16
17
18
19
20
21
22
D15
D24
D26
D30
D36
D40
D50
D64
D75
D100
Eleutheroembryo
Juvenile
Characteristics
Figure
Freshly spawned egg
Neural plate
Onset somatogenesis
Brain regionalisation
Early trunk-tail
Neural anlage
Pectoral fin bud
Pre hatching
Hatching
Free embryo
Jaw and branchial placodes
Initiation of barbels
Ventral fin bud and swim
bladder
Late eleutheroembryo
Juvenile 1
Juvenile 2
Juvenile 3
Juvenile 4
Juvenile 5
Juvenile 6
Juvenile 7
Juvenile 8
52a
52b-c
53a-c
54a-b
55a-d
56a-d
57a-b
58a-b
58c-d
59a
59b
61a
61b
61a
63b
63c
63d
64a
64b
64c
64d
64e
80
4 DISCUSSION
4.1 Overview of environmental triggers of gonad development
4.1.1 Family Mormyridae
The environmental factors (decreasing conductivity, constant water level and slight
variation of temperature) applied in the experiment with one female and three males
clearly
showed
a
significant
influence
on
gonad
maturation
in
Petrocephalus soudanensis. The female has shown a largely swollen abdomen. This is
in accordance with Kirschbaum (2006) and similar to other investigated mormyrids
(Schugardt and Kirschbaum, 2004). The pair also established territories in the tank,
while conductivity was lowered; however, no spawning occurred. This could be due to
stress factors such as insufficient food during the breeding period or a too small
breeding tank. Kirschbaum (1992) stated that stress factors prevent the completion of
reproductive behaviour. The P. soudanensis used in this experiment had been used in
Kirschbaum’s experiments (2006) with successful breeding.
In the first breeding experiment with Petrocephalus catostoma (four males and female),
a pair established its territory in the tank, however, no spawning was observed. Similar
to the second breeding experiment with a female and two males, neither courtship
behaviour nor spawning was observed. The influence of environmental factors
decreasing conductivity, constant water level and slight variation of temperature on
gonad development therefore remains unclear in this species. The breeding experiments
failed in P. catostoma presumably due to the age of the specimens. Since the exact age
was unknown, it is possible that they had already reached the senescent period. In fact,
there was a significant decrease in total weight of the specimens within a-9-month
experimental period.
4.1.2 Family Notopteridae
In the two breeding experiments with Xenomystus nigri, there is no clear influence of
environmental factors (decreasing conductivity, constant water level and slight variation
of temperature) on gonad maturation. In this study, two females showed gonad
maturation in the breeding group I (9 females, 7 males). Three females also showed
swollen bellies from the breeding group II comprising nine females and male. Thus,
there was a mixed reaction since swollen bellies were found during rainy
(decreasing conductivity) and dry season (increasing conductivity) in both experiments.
No spawning activity occurred in both experiments. Courtship behaviour occurred
sporadically in these two breeding groups. Apparently the small size of the breeding
81
tank and missing of a preferable spawning substrate could be the main reason for the
failing of the breeding experiments. Nyonje (2006) stated that the three environmental
factors imitation of rain, decrease conductivity and increase water level do not regulate
gonad maturation in X. nigri.
In the dissected gonads of X. nigri, there were no ovulated eggs found, although some
females’ gonads contained oocytes in maturation stage (MC: 7.70 %; 7.63 % of
breeding group I and MC: 8.3 % of breeding group II). The ovaries were filled with
different stages of oocytes indicating fractional spawner.
In order to collect data for ontogeny development, trials of artificial reproduction have
been applied to the selected pairs with swollen bellies. Intense courtship behaviour was
still seen after the hormonal treatment, even though attempts of hormonal injection
failed as no spawning occurred. In most cases of fractional spawners, artificial
reproduction is difficult to perform.
Similar to X. nigri, there is no obvious reaction to the environmental factors (decreasing
conductivity, constant water level and slight variation of temperature) in
Notopterus notopterus. A female of the breeding experiment I (two females and male),
indeed showed a swollen belly after the onset of decreasing conductivity. Intense
courtship behaviour observed until the first spawning took place after six months
experimental period. In the breeding group II comprising a pair of N. notopterus,
swollen bellies as indication for gonad maturation and courtship behaviour were often
seen at high levels of conductivity. The influence of environmental factors on the gonad
maturation and courtship behaviour in N. notopterus are therefore unclear. Apparently
endogenous factors play an important role predominantly in regulating gonad
maturation and in provoking courtship behaviour in this species. A study of Weitkamp
(2005) on N. notopterus revealed that none of the environmental factors tested (i.e
conductivity, temperature and water level) had significant influence on gonad
maturation.
4.1.3 Family Pantodontidae
Two experiments were conducted in Pantodon buchholzi regarding environmental
factors: constant water level, decreasing conductivity and slight variation of
temperature. In the breeding group I containing seven males and five females, no
spawning occurred even though a pair showed clear signs of courtship behaviour for
several days during decreasing conductivity. In the breeding group II with one male and
two females, neither gonad maturation nor courtship behaviour was observed during the
82
experimental period. This could be due to wrong feeding or maybe the applied
parameters were not appropriate to induce the gonad maturation in this species.
Conductivity was drastically decreased a few times in the three additional breeding
experiments with P. buchholzi, comprising breeding group combination (BGC) I (1♀,
1♂), BGC II (5♀, 4♂), and BGC III (4♀, 5♂). Neither a swollen belly nor courtship
behavior was observed throughout these trials. It seems that environmental factors do
not regulate gonad maturation in P. buchholzi. Britz (2004) reported that after several
drastic water changes gonad maturation in P. buchholzi could be induced. Apparently
maturation is controlled endogenously and the environmental factors only provoke
ovulation in P. buchholzi, as found in Polypterus senegalus (Kirschbaum, 1992; 1995b;
Yanwirsal, 2007).
4.1.4 Family Osteoglossidae
Based on the results of fieldwork in Colombia, while collecting the samples, this has
shown that the rainy season (January to May) is the main spawning season in
Osteoglossum bicirrhosum. This is confirmed by the experience of the local farmers that
in general the onset of rainy season is often used as a definite sign to indicate the
spawning season in O. bicirrhosum. This is in accordance with Argumedo (2005):
breeding season occurs during the rainy season (in late November to early of July). In
contrast to other osteoglossids, spawning mainly occurs prior to the onset of the rainy
season in Scleropages formosus (Scott and Fuller, 1976; Suleiman, 2003).
4.2 Modes of reproduction in Notopterus notopterus and Osteoglossum bicirrhosum
with comparison to other osteoglossomorphs
4.2.1 Reproductive guilds
This study confirms that Notopterus notopterus is a substrate spawner with a semitransparent yellowish egg envelope. This is in accordance with Mookerjee and
Mazumdar (1946), Ong (1965), Axelrod and Burgess (1981), and Pinxteren (1974).
Substrate spawning is also performed by Chitala chitala (Pinxteren, 1974; Axelrod and
Burgess, 1981).
This study also verifies that Osteoglossum bicirrhosum is a mouth breeder
(Ungar, 1993; Argumedo, 2005) with a large (Wolfsheimer, 1964) translucent envelope.
All the representatives of the genera Scleropages are mouth breeders (Merrick and
Green, 1982; Lake and Midgley, 1970; Azuma, 1992; Brown, 1995). Nest building
behaviour is seen in male mormyrids of Pollimyrus isidori and Pollimyrus adspersus
(Diedhiou et al., 2007a); and in the two osteoglossids Heterotis niloticus
83
(Budgett, 1901a; Svensson, 1933; Johnels, 1954; Daget, 1957) and Arapaima gigas
(Fontanele, 1948, 1952; Lüling, 1964, 1969; Neves, 1998). Budgett (1901a) and
Svensson (1933) reported that only the species Gymnarchus niloticus built large floating
nest.
4.2.2 Spawning time
Spawning in N. notopterus occurs mostly during day time. Friese (1980) also reported
that spawning in N. notopterus tends to occur in the early morning. In contrast, Smith
(1933) and Pinxteren (1974) stated that spawning in N. notopterus occurs mainly at
night. It seems that spawning in N. notopterus may occur at any time. In the Silver
Arowana, O. bicirrhosum spawning takes place at night and in the early morning.
Spawning in S. leichardtii apparently takes place at night (Merrick and Schmida, 1984),
similar to Pollimyrus isidori (Schugardt and Kirschbaum, 2006; Diedhiou et al., 2007b),
Mormyrus rume probocirostris
(Schugardt
and
Kirschbaum,
2004)
and
Campylomormyrus cassaicus (Schugardt and Kirschbaum, 2002), Pantodon buchholzi
(Britz, 2004) and Petrocephalus soudanensis (Kirschbaum, 2006).
4.2.3 Left gonad
This study verifies that O. bicirrhosum and N. notopterus possess a single gonad located
on the left side of the body, as observed also in Xenomystus nigri and
Osteoglossum ferreirai. Therefore it confirms the statement of Nyonje (2006) and
Argumedo (2005, 2009) concerning the presence of one gonad in X. nigri,
O. bicirrhosum and O. ferreirai. A single gonad is also present in
Scleropages leichardtii (Merrick and Schmida, 1984; Merrick et al., 1983; Lake and
Midgley,
1970),
Heterotis niloticus
(Moreau,
1974),
Arapaima gigas
(Fontanele, 1948; Lüling, 1964), Chitala ornata (Smith, 1933) and in three mormyrids
Hyperopisus bebe and Mormyrus kannume (Nawar, 1959; Scott, 1973) and also in
Hippopotamyrus pictus (Blake, 1977). In addition, this study also confirms that
Pantodon buchholzi possesses both ovaries. This is in accordance with Nysten (1962).
The presence of both ovaries was also found in the Mooneye Hiodon tergisus (Glenn
and Williams, 1976). Britz (2004) concluded that species with paired ovaries possess
the plesiomorphic and those with only one ovary the apomorphic character.
4.2.4 Fractional spawner
The ovary of O. bicirrhosum and N. notopterus is filled with various stages of oocytes
comprising stage I (primary growth) to stage IV (maturation stage) (based on Wallace
and Selman, 1981). This indicates that both species are fractional spawners. Azadi et al.
84
(1995) showed that the ovary of N. notopterus was filled with three different stages (I to
III) of oocytes. Similar to what Weitkamp (2005) found, the ovary of N. notopterus
contained different stages of gonad maturation. Fractional spawning was also observed
in Pollimyrus isidori (Kirschbaum, 1987; Kirschbaum and Schugardt, 1995); in
Mormyrus rume probocirostris (Schugardt and Kirschbaum, 2004); in fishes of the
Paramormyrops magnostipes-complex (Nguyen, 2011); and in Xenomystus nigri (this
study; Nyonje, 2006).
4.2.5 Parental care
This study proved that N. notopterus shows parental care performed by the male as
reported by other authors (Pinxteren, 1974; Smith, 1933; Axelrod and Burgess, 1981;
Friese, 1980). Both Chitala ornata and Chitala chitala take parental care of their
adhesive eggs. The male guards the eggs attached to different substrates (Smith, 1933;
Southwell and Prashad, 1919). Trittelvitz (1986) and Sieraad (1999) assumed that there
was no parental care in Xenomystus nigri.
In mormyrids, Pollimyrus isidori and P. adspersus are known to show behaviour of
parental care (Kirschbaum and Schugardt, 2002; Diedhiou et al., 2007a), as well as in
Stomatorhinus (Heymer and Harder, 1975). No signs of parental care was reported
neither for Campylomormyrus cassaicus nor Hippopotamyrus pictus (Kirschbaum and
Schugardt, 2002); nor for Paramormyrops magnostipes-complex (Nguyen, 2011).
Pantodon buchholzi also does not perform parental care (Britz, 2004; Siegl, 1914;
Schreitmüller, 1936; Vriends, 1978)
The male of O. bicirrhosum carries the eggs, embryos and juveniles for about two
months in its mouth. According to several authors (Argumedo, 2005; Merrick and
Green, 1982; Azuma, 1992; Brown, 1995; Takeshita, 1973; Scott and Fuller, 1999;
Dawes et al., 1999) mouth breeding in the family Osteoglossidae: O. bicirrhosum,
S. leichardtii, S. jardini is performed by the male. A recent paper by Schaefer (2010)
reported that female of Scleropages was occasionally found carrying eggs, though this
statement was denied by the breeders.
4.2.6 Egg adhesiveness
The adhesive eggs from N. notopterus were always attached to a substrat, i.e. either to a
rock, a huge/flat stone or to a filter. This is in line with Srivastava et al. (2012), Friese
(1980), Axelrod and Burgess (1981), and Pinxteren (1972). Adhesive eggs are also
found in other notopterid species such as Chitala ornata (Smith, 1933) and in five
mormyrids: Hippopotamyrus pictus, Campylomormyrus cassaicus, C. phantasticus, and
Mormyrus rume probocirostris (Kirschbaum and Schugardt, 2002) and in the fishes of
85
Paramormyrops magnostipes-complex (Nguyen, 2011). Daget (1957) also reported that
the eggs of Heterotis niloticus sink to the bottom of the nest, where they adhere to each
other, forming a large mass of eggs.
This study reveals for the first time that the eggs of O. bicirrhosum are non-adhesive.
Non-adhesive eggs can also be found in other osteoglossomorphs, such as
Pollimyrus isidori and P. adspersus (Diedhiou et al., 2007a). Concerning
Pantodon buchholzi (Britz, 2004) it is reported to produce buoyant eggs with some
large oil globules. The eggs of Hiodon alosoides are semi pelagic eggs due to its single
large oil globule (Battle and Sprules, 1960). Wallus (1990) reported that the eggs of
H. tergisus and H. alosoides are non-adhesive and buoyant/semi buoyant.
4.2.7 Egg numbers per spawning
In this study the number of eggs per spawning in Notopterus notopterus varies from
15-225 eggs. Some previous authors reported number of eggs per spawning in between,
namely 50 eggs (Ong, 1965), up to 90 eggs (Pinxteren, 1974) and around 30–100 eggs
(Axelrod and Burgess, 1981). A higher number of eggs compared to N. notopterus was
found in Chitala chitala, 300-500 eggs (Southwell and Prashad, 1919) and up to 3500 in
C. ornata (Smith, 1933). In Xenomystus nigri, the number of eggs per spawning varies
from 40–60 (Trittelvitz, 1986).
The male Osteoglossum bicirrhosum may carry around 23-220 eggs. This study
presents higher numbers of eggs per spawning in O. bicirrhosum in comparison to
Maupin (1967), Ungar (1993) and Wolfsheimer (1964), who reported that
O. bicirrhosum spawned between 40, 135 and 150 eggs. Less number of eggs per
spawning was also found in other osteoglossids, around 30–80 in Scleropages formosus
(Azuma, 1992), around 30–110 in S. leichardtii (Allen et al., 2002), and 11.000 eggs in
Arapaima gigas (Neves, 1998).
There is a wide variety of the quantity of eggs per spawning in other
osteoglossomorphs. Regarding mormyrid species, Pollimyrus isidori lays 30–200 eggs
(Kirschbaum, 1987; Kirschbaum and Schugardt, 1995); P. adspersus around 28–215
(Diedhiou et al., 2007a); Petrocephalus soudanensis 180–283; C. phantasticus about
250–400; Mormyrus rume probocirostris 128–728, and Hippopotamyrus pictus lays
circa 66–772; Campylomormyrus cassaicus 121–1662 (Kirschbaum and Schugardt,
2002); and about 1000 eggs in Gymnarchus niloticus (Budgett, 1901a; Svensson, 1933).
86
4.2.8 Egg size
In this study, the size of freshly collected eggs from Osteoglossum bicirrhosum mainly
measured 12 mm. This differs from the two previous authors Wolfsheimer (1964) and
Aragao (1981) showed that the eggs of O. bicirrhosum are 16 mm and 13 mm large. In
other osteoglossids, such as Scleropages formosus, the newly spawned eggs measured
19 mm in diameter (Shigeru et al., 1999), around 15–18 mm (Azuma, 1992), reached up
to 10 mm in S. leichardtii (Lake and Midgley, 1970), and 3 mm (Menezes, 1951) in
Arapaima gigas. The egg size varies from 2.5 mm (Budgett, 1901a) and circa
2.5-2.8 mm (Daget, 1957) in Heterotis niloticus; 2.8 mm (Fontanele, 1952), and 4.2 mm
(Fontanele, 1948).
The newly spawned eggs of N. notopterus in this study were 3.8–4 mm in diameter.
This is in accordance with Axelrod and Burgess (1981), Svensson (1933), and a recent
study of Srivastava et al. (2012). In contrast to what Friese (1980) and Azadi et al.
(1995) reported, the eggs of N. notopterus are 3.5 mm and 3.10 mm in size.
Except for the eggs of Gymnarchus niloticus which measure 10 mm in diameter
(Budgett, 1901b), it seems that the size of the eggs in notopterids and osteoglossids are
larger than in other osteoglossomorphs. As seen in the egg size of mormyrids such as
1.8 mm in Hyperopisus bebe (Johnels, 1954) and Petrocephalus soudanensis
(Kirschbaum, 2006); 2 mm in Paramormyrops magnostipes-complex (Nguyen, 2011),
Pollimyrus isidori (Kirschbaum, 1987; Diedhiou et al., 2007), Marcusenius mento
(Boulenger, 1890; Schugardt and Kirschbaum, 2006), Campylomormyrus tamandua
(Günther, 1864), and Campylomormyrus cassaicus (Schugardt and Kirschbaum, 1998);
3 mm in Xenomystus nigri (Trittelvitz, 1986), Campylomormyrus phantasticus and
Hippotamyrus pictus (Kirschbaum and Schugardt, 2002).
Concerning Pantodon buchholzi the egg size measure 2 mm (Mohn, 1976a), circa
2.2-2.4 mm (Britz, 2004). In hiodotid species, egg of Hiodon tergisus and H. alosoides
measures 4 mm and 3.3-3.9 mm in diameter (Wallus, 1990).
4.2.9 Hatching
In this study it has been shown that N. notopterus hatched at 7–8 days after spawning at
a of temperature of 27 °C This is a bit different to what other authors found: 5–6 days
after spawning at 26-28 °C (Srivastava et al., 2012); 5–6 days after spawning
(Pinxteren, 1974); after 7 days of spawning (Axelrod and Burgess, 1981); in 6 days
after spawning at 28 °C (Friese, 1980). In Chitala chitala hatching occurred around five
days after spawning (Hossain, 1999; Radheyshyam and Sarangi, 2005).
87
Hatching in Osteoglossum bicirrhosum occurs around 7 days at 28 °C after being
released from the male’s mouth. In other osteoglossids, hatching occurs in 7-14 days in
Scleropages leichardtii (Lake, 1971; 1978), 12 days after spawning in S. formosus
(Azuma, 1992), five days (Neves, 1998) and around 10–14 days in Arapaima gigas
(Lake and Midgley, 1970).
In Heterotis niloticus (Budgett, 1901a) hatching occurred around two days after
spawning; after three days in fishes of the Paramormyrops magnostipes-complex
(Nguyen, 2011), Petrocephalus soudanensis (Kirschbaum, 2006; Kirschbaum and
Schugardt, 2002), Pollimyrus isidori (Kirschbaum, 1987; Kirschbaum and Schugardt,
1995; Diedhiou et al., 2007) and Mormyrus rume probocirostris (Kirschbaum and
Schugardt, 1995); and after three days (Britz, 2004; Schreitmüller, 1936) and 4–6 days
in Pantodon buchholzi (Siegl, 1914). In Hiodon alosoides hatching occurs about
14 days after spawning (Wallus, 1990).
However the developmental stage at hatching is influenced by environmental factor
such as temperature and oxygen conditions (Hamor and Garside 1979;
Peňáz et al. 1983; Heming, 1982).
4.2.10 Size of free embryos at hatching
In this study, free embryos of N. notopterus, directly after hatching, measured 10.5 mm
in total length, which however, is slightly longer to what Axelrod and Burgess (1981)
found, according to whom newly hatched N. notopterus measured 7 mm. Newly
hatched embryos of N. notopterus measured 8.3 mm in total length (Mookerjee and
Mazumdar, 1946) and 8.0±0.5 mm long (Srivastava et al., 2012). In other notopterids,
free embryos of Chitala chitala measured 13.8 mm (Southwell and Prashad, 1919) and
12 mm in C. ornata (Smith, 1933).
The size of the free embryos in Osteoglossum bicirrhosum in this study measured
16 mm in total length at hatching which occurred 7 days after being released from the
mouth of male’s parent. Argumedo (2005) reported that the free embryos of
O. bicirrhosum are 17-20 mm long after hatching.
S. leichhardtii’s large-sized embryos measured 36 mm (Lake and Midgley, 1970) and
15 mm at hatching (Merrick and Green, 1982). Free embryos in Heterotis niloticus
measured in total length of 7.5 mm (Daget, 1957), whereas 11.6 mm long in
Arapaima gigas (Neves, 1998; Fontanele, 1948, 1952);
A variable hatching size of embryos (in total length) was also reported in other
osteoglossomorphs: 3.4 mm in Pollimyrus isidori (Diedhiou et al., 2007b) and
88
P. adspersus
(Diedhiou
et
al.,
2007a);
3.5–4 mm
in
the
fishes
of
Paramormyrops magnostipes-complex
(Nguyen, 2011)
and
4.2 mm
in
Campylomormyrus cassaicus (Schugardt and Kirschbaum, 1998), 7 mm in
Hippopotamyrus pictus (Kirschbaum and Schugardt, 2002); around 7.2 mm in
Petrocephalus soudanensis (estimated from Fig. 3 in Kirschbaum, 2006); 4.2–4.6 mm
in Pantodon buchholzi at 29 °C (Britz, 2004); ca. 7 mm in Hiodon tergisus
(Wallus, 1990) and 7.2-7.6 mm in H. alosoides (Battle and Sprules, 1960).
4.2.11 Onset of exogenous feeding
Exogenous feeding was observed for the first time in N. notopterus larva after 17 days
with 16.2 mm in total length. Similar to Chitala ornata, exogenous feeding started once
larvae were 13–18 mm long and 16–17 days old (Smith, 1933). The resorption of yolk
sac in N. notopterus, observed around 18 days after spawning with 15.3 mm in total
length (Mookerjee and Mazumdar, 1946) and at the age of 22 days (Axelrod and
Burgess, 1981), can be categorized as the onset of exogenous feeding. O. bicirrhosum
started mixed feeding on the 26th day with 38.2 mm in total length. Shigeru et al. (1999)
reported that 64 days after spawning the fry of S. formosus began to leave the male’s
mouth, but only for a short time, and after 111 days with a total length of 62 mm it
proceeded to do so permanently. 60 days after spawning the fry for the first time swam
free for a longer period of time, though they still stayed close to the male’s mouth
(Azuma, 1992).
Regarding other osteoglossomorphs, reports of the emergence of exogenous feeding are:
3 days (Kirschbaum and Schugardt, 2002) and 4 days (Kirschbaum, 1994; Diedhiou et
al., 2007) in Pollimyrus isidori; 8 days after hatching in Mormyrus rume probocirostris
(Kirschbaum and Schugardt, 1995), Campylomormyrus cassaicus (Schugardt and
Kirschbaum, 1998), Hippopotamyrus pictus (Kirschbaum and Schugardt, 2002),
Petrocephalus soudanensis (Kirschbaum and Schugardt, 2002); in the fishes of
Paramormyrops magnostipes-complex
(Nguyen, 2011),
and
4–5
days
in
Pantodon buchholzi (Britz, 2004).
The onset of exogenous feeding is depending on the egg size of the species. It seems
that the larger the egg size, the later the onset of exogenous feeding begins.
89
4.3 Characteristic of the egg envelope, eye pigmentation, melanophore pattern
and fins development in Notopterus notopterus and Osteoglossum bicirrhosum with
comparison to other osteoglossomorphs
The egg of Notopterus notopterus has many external ridges which are centred around
the single micropyle located at the animal pole. Mookerjee and Marzumdar (1946) also
stated that eggs of N. notopterus show a “groove like line, radiating from the
micropyle”. Based on the four classifications of micropyle described by Kunz (2004,
modified from Riehl, 1991), the micropyle of N. notopterus belongs to type number 1:
the eggs surface displays a spiralling pattern of ridges partially ending in the micropylar
region (Riehl and Kokoschka, 1993). The pattern of the furrows on the surface of these
species is radial, running from the animal to the vegetal pole (Kunz, 2004). The egg of
the catfish Sturisoma aureum of the family Loricariidae (Riehl and Patzner, 1991) and
the cyprinid Barbus conchonius of the family Cyprinidae (Amanze and Iyengar, 1990)
also possess a micropyle with grooves and ridges directed toward the micropylar canal.
The chorion in two other osteoglossomorphs Pollimyrus isidori (Diedhiou et al., 2007b)
and Pantodon buchholzi (Britz, 2004) appears smooth at binocular magnification.
Eye pigmentation in both N. notopterus and O. bicirrhosum commence within the egg
envelope once five or six days old. It is proven by Argumedo (2005) that at stage I,
10.5 mm long embryos O. bicirrhosum show first signs of pigmented eyes inside of the
egg envelope. In contrast to Pollimyrus isidori (Diedhiou et al., 2007b), first indicators
of pigmented eyes emerged on the fourth day after hatching in P. adspersus (Diedhiou
et al., 2007a) and Petrocephalus soudanensis (Kirschbaum, 2006) and a few hours after
hatching in fishes of the Paramormyrops magnostipes-complex (Nguyen, 2011).
The onset of melanophore formation in N. notopterus emerges around the forehead prior
to hatching, at the age of five days. In contrast to this, a massive black pigmentation
first appears to be covering the brain region of O. bicirrhosum on the first day after
hatching. This was also found by Argumedo (2005): black melanophores were first seen
after hatching, in stage II fry in 17–20 mm long individuals.
In mormyrids, the first melanophores appear in Pollimyrus isidori the day after hatching
in the dorsal head epidermis (Diedhiou et al., 2007). In fishes of the
Paramormyrops magnostipes-complex, black melanophores developed one day after
hatching on the top of the head (Nguyen, 2011). In Pantodon buchholzi, black
melanophores were observed for the first time on the head areas, on the body and on a
reticulate pattern on the dorsal region of the yolk-sac (Britz, 2004). The free embryos of
Hiodon alosoides lack melanophores completely, although occasionally melanophores
could appear on the mid-dorsal surface (Battles and Sprules, 1960). As for
90
Hiodon tergisus, Synder and Douglas (1978) reported that some branched melanophores
appeared a few days after hatching, distributed on the ventral half of the yolk sac.
Some colouring is present in N. notopterus, especially starting at the age of 52 days
until 92 days. In contrast, Pantodon buchholzi shows a unique coloration in the larval
period only (Britz, 2004).
The development of pectoral, dorsal, anal, caudal, and fins already starts prior to the
onset of exogenous feeding in Notopterus notopterus. This is similar to
Pollimyrus isidori, P. adspersus, Mormyrus rume probocirostris (Diedhiou et al.,
2007b) and also Pantodon buchholzi (Britz, 2004). In three other mormyrids
Campylomormyrus tamandua, Petrocephalus soudanensis and Hippopotamyrus pictus:
the demarcation of the dorsal and anal fins emerges later, during the larval period
(Diedhiou et al., 2007b). Similar to N. notopterus and other forementioned mormyrid
species, the development of fins in Osteoglossum bicirrhosum, starts with pectoral fins,
followed by dorsal, anal, caudal and at last with the pelvic fins.
4.4 Comparison of periods and phases in development of Notopterus notopterus
and Osteoglossum bicirrhosum
Fig. 65. Comparison of periods and phases in the development of Notopterus notopterus and
Osteoglossum bicirrhosum (based on terminology of Balon, 1975). Neural plate (NP) as the beginning of
the time scale.
91
In this study, a nearly complete series of development stage in Notopterus notopterus
and Osteoglossum bicirrhosum is described for the first time. Figure 65 illustrates the
comparative development as divides into periods and phases in both N. notopterus and
O. bicirrhosum.
Neurulation, the externally well visible development of the neural plate is used as the
standard onset of the time scale in both development courses for comparative purpose.
The neural plate (NP) stage in N. notopterus sets in at 29 hours after spawning and is
observed in O. bicirrhosum at 8 hours after collecting time.
4.4.1 Periods
There are three periods described in Figure 65: the embryonic period, the larval period
and the juvenile period. In N. notopterus compared to O. bicirhosum, the embryonic
period lasts shorter and the onset of juvenile period starts much delayed. The larval
period lasts long before shifting to the juvenile period in N. notopterus. The larval
period in N. notopterus is characterized by obliteration of the embryonic fin fold, a fully
resorbed yolk sac, a distinct larval coloration and the onset of exogenous feeding. The
temporary embryonic fin fold appears after 95 hours and lasts until day 13 in
N. notopterus. The larval period is then followed by the juvenile period starting on day
51 and lasting until the age of 18 months.
The larval period is absent in O. bicirrhosum since none of the temporary organs were
found. This is in contrast to Argumedo (2005) who mentioned the existence of the larval
period in O. bicirrhosum, probably by implicitly using another definition of a larval
phase.
The start of the juvenile period is characterized by the the onset of mixed feeding in
O. bicirrhosum, which was observed at day 26. At the age of 36 days with a total length
of 45 mm, the juveniles of O. bicirrhosum have already accomplished the formation of
the definite organs such as well-developed pelvic fins, barbels, and the onset of scale
formation, including the almost entirely absorbed yolk sac and the completed juvenile
colouring. At this point the juveniles are already capable of leaving the parent’s mouth
and try to swim individually in their natural habitat. Around day 100, O. bicirrhosum
reaches a length of 125 mm with the fully absorbed yolk-sac and the juveniles resemble
young adults. The juvenile period in O. bicirrhosum lasts for a very long time until it
turns into an adult. O. bicirrhosum is fully sexually mature from the age of 28 months
on.
92
The absence of the larval period in O. bicirrhosum leads to a conclusion that this
species undergoes a direct development, while N. notopterus undergoes an indirect
development including the larval period.
4.4.2 Phases
The embryonic period consists of two phases: the embryonic phase and the
eleutheroembryonic phase. The embryonic phase indicates the development within the
egg envelope until hatching occurs. This phase lasts nearly similarly long in both
species.
The eleutheroembryonic phase in N. notopterus commences shortly after hatching and
lasts until the onset of exogenous feeding. While, the eleutheroembryonic phase in
O. bicirrhosum starts after hatching and lasts until the first mixed feeding while the yolk
sac is still attached. The eleutheroembryonic phase in O. bicirrhosum is almost twice as
long as in N. notopterus.
4.4.3 General comments
A large yolk sac in O. bicirrhosum, allows it to protect its offspring during and after
mouth breeding, especially providing massive food supply endogenously. Producing
large yolky eggs, the embryos of O. bicirrhosum tend to stay longer inside the egg
envelope which enables further protection by parental care.
Possessing a large yolk sac allows the embryos to instantly structure the whole suite of
permanent organs without being required to modify any temporal larval structures
(Balon, 1984b). Oppenheimer (1970) stated that, the greater the amount of yolk in the
eggs, the less likely parental care will be continued after the fry are released.
Mixed feeding in O. bicirrhosum, referring to exogenous feeding alongside with the
yolk-sac still attached. This is in accordance with Balon (1984b) who mentioned that
often the initial action of a juvenile period in direct developing organisms is categorized
by mixed feeding of varying duration. Mixed feeding is delayed for around 10 days in
O. bicirrhosum compared to N. notopterus. Elimination of the vulnerable larva and
metamorphosis may facilitate direct development into a juvenile that is relatively
advanced at the first oral feeding (Balon, 1999). Mixed feeding also occurs in the mouth
breeding Cyphotilapia frontosa (Cichlidae) starting with free embryos until early
juvenile (Balon, 1999). In a cichlid species Labeotropheus, the eleutheroembryo
develops directly into a juvenile without metamorphic stages, forming advanced
structures like fins, skeleton and pigments at a time when a large yolk sac is still present
93
(Balon, 1977). As found in most species, the juveniles will soon live in shallow nursery
habitats, which are unreachable to larger piscine predators (Balon, 1999).
Different models of life history for fish were proposed by some authors. Winemuller
(2005) illustrated a triangular model: 1) opportunistic – short generation times and small
body size, producing a large number of eggs with no parental care (comparable to rstrategist), 2) periodic species – long-living individuals producing lots of eggs, but
provide no parental care, and 3) equilibrium species – living fairly long with few
offspring and provide parental care (comparable to K-strategist). Based on his concept
concequently N. notopterus belongs to the opportunistic model and equilibrium model
matches O. bicirrhosum.
Egg size in Notopterus is also shown to be not that diminutive as in typical textbook
examples of r-strategic reproduction, since this species produces medium-sized eggs
with parental care. N. notopterus therefore is classified as an intermediate species with
respect to interpretation of a reproductive strategy, while O. bicirrhosum is classified as
a K-strategic. Rather, each species apparently evolves a unique suite of adaptive
characters, in order to survive in their environment and to protect their offspring. This is
well demonstrated in the wide range of reproductive styles among Osteoglossomorpha –
and the terminology and stage definitions of early ontogenetic development in fish
established by Balon (1975a, b) is well suited to characterize these various reproductive
styles in the light of adaptiveness.
94
5 CONCLUSION
This study essentially contributes to the knowledge of the reproduction and the
ontogenetic development in two species of Osteoglossomorpha belonging to two
families: Notopterus notopterus (Notopteridae) and Osteoglossum bicirrhosum
(Osteoglossidae).
The only comparable in-depth study is available in Mormyridae (Pollimyrus isidori).
For the first time the ontogenetic development is described in detail and fairly complete
in the mouth breeder Osteoglossum bicirrhosum. This ontogeny is characterized by the
absence of a larval period and thus representing direct development, whereas
Notopterus notopterus represents a more typical indirect development.
A broader comparison of the various reproductive styles in Osteoglossomorpha, also
based on ontogenetic data of previous authors, characterizes Osteoglossum bicirrhosum
as “k-strategic” and Notopterus notopterus as “intermediate” with respect to
interpretation of a reproductive strategy.
Thus this study has considerably increased our understanding of reproduction and
ontogenetic development in the basal and very diverse taxon Osteoglossomorpha
95
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104
ACKNOWLEDGEMENTS
Al-hamdu lillahi rabbil 'alamin
I would like to express my sincerest gratitude to Prof. Dr. Frank Kirschbaum for his
guidance, motivation, unconditional help, immense patience and recommendations in
many ways. I also thank his spouse Mrs. Yvonne Kirschbaum for her help and moral
support.
I would also like to extend my deepest gratitude to Dr. Peter Bartsch for his guidance,
cooperation, motivation, and encouragement.
My deepest gratitude goes to Mr. Yousef Jameel for his valuable financial support.
I thank Prof. Dr. Norbert Walz and Prof. Dr. Ralph Tiedemann for their
cooperation.
I am also grateful for the kind support of Mr. Christian Hoffmann and Dr. Uta
Hoffmann.
My sincere gratitude goes to H. Yan Wirsal, Hj. Sumiarti, Madonna and Holy Fika
for their unconditional love, trust and prayers.
My deepest gratitude also goes to Rafael Luty and family for their continuous support,
attention, motivation, and faith.
Special thanks go to Hartmut Höft, Wolfgang Bernau, Suzanne Grübel,
Petra Grimm, Annett Billepp and Jutta Zeller for their guidance and help in the
laboratory.
I thank H. E. Dr. Eddy Pratomo the Ambassador of the Republic of Indonesia in
Berlin and Mr. Michael Manufandu the former Ambassador of the Republic of
Indonesia in Bogota, for their kind support, attention and motivation.
I gratefully acknowledge Mr. Subagia Made, Ms. Abigail Sihotang, Mr. Agus
Prabawa and all staff of Indonesian Embassy in Bogota for their tremendous help
and encouragement.
I gratefully thank Eric Giovanny Argumedo Trilleras, David Garcia, Joell, and their
families for their valuable cooperation and kindness.
105
My huge gratitude goes to Mrs. Nancy Quintero Ramirez Director of ACUICA
(Asociación de Acuicultores del Caqueta), Jose Alexander Lopez, Christian
Fernando Erazo Acosta, Hugo Rojas, Gladys and all the farmers for their
contribution in providing investigated materials and their guidance at field work.
I thank Mr. Alexander Velasquez Manager of Museo de Historia Natural and
Alexander Claros Diaz Manager of Biology Laboratory of Universidad de la
Amazonia of Florencia for their valuable cooperation in providing documentation
devices.
Special thanks to Julio Hernan Lopez and Tania Ramsa for their kind assistance in
the laboratory in Florencia.
My sincerest gratitude goes to Mrs. Fanny Quesada, Virgilio Ibarro Garcia and all
members of CremaPan family for their tremendous help, support and kindness during
my hard times in Colombia.
Special
thanks
to
Yorcelys Cruz,
Hany
Abdelkawi,
Dr. Reza Fard,
Dr. Farzana Anjum, Dr. Salif Diedhiou, Dr. Betty Nyonje, Jostein Kraakas,
M. Abas Ridwan, and Dr. Azwir Gusrialdi for their encouragement and kind support.
I thank the members of my defense committee for their precious time to read my
dissertation and for their insightful comments.
Last but not least, I would like to thank all my friends from all over the world. Without
their continuous support and friendship I could not have gotten this far.
106
SELBSTÄNDIGKEITSERKLÄRUNG
Hiermit erkläre ich, die Dissertation selbständig und nur unter Verwendung der
angegebenen Hilfen und Hilfsmittel angefertigt zu haben.
Ich versichere, dass die Dissertation bisher weder in Teilen noch als Ganzes einem
Promotionsverfahren zugrunde lag.
Ich habe mich nicht anderwärts als Doktorand beworben und besitze keinen
entsprechenden Doktorgrad.
Ich erkläre die Kenntnisnahme der dem Verfahren zugrunde liegenden
Promotionsordnung
der
Landwirtschaftlich-Gärtnerischen
Fakultät
der
Humboldt-Universität zu Berlin.
Berlin, den 7. Januar 2013
Honesty Yanwirsal